15211878, 0, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/bies.202200123 by Readcube (Labtiva Inc.), Wiley Online Library on [16/01/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License Received: 25 June 2022 Revised: 14 October 2022 Accepted: 5 December 2022 DOI: 10.1002/bies.202200123 I D E A S & S P E C U L AT I O N S Insights & Perspectives Three dimensions of thermolabile sex determination Paul D. Waters1 Jennifer A. Marshall Graves2,3 Sarah L. Whiteley3 Arthur Georges3 Aurora Ruiz-Herrera4,5 1 Faculty of Science, School of Biotechnology and Biomolecular Science, UNSW Sydney, Abstract Sydney, NSW, Australia The molecular mechanism of temperature-dependent sex determination (TSD) is a 2 Department of Environment and Genetics, La long-standing mystery. How is the thermal signal sensed, captured and transduced to Trobe University, Bundoora, Australia 3 Institute for Applied Ecology, University of regulate key sex genes? Although there is compelling evidence for pathways via which Canberra, Canberra, Australia cells capture the temperature signal, there is no known mechanism by which cells 4 Departament de Biologia Cellular, Fisiologia i transduce those thermal signals to affect gene expression. Here we propose a novel Immunologia, Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain hypothesis we call 3D-TSD (the three dimensions of thermolabile sex determination). 5 Genome Integrity and Instability Group, We postulate that the genome has capacity to remodel in response to temperature Institut de Biotecnologia i Biomedicina, by changing 3D chromatin conformation, perhaps via temperature-sensitive transcrip- Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain tional condensates. This could rewire enhancer–promoter interactions to alter the expression of key sex-determining genes. This hypothesis can accommodate mono- Correspondence Paul D. Waters, Faculty of Science, School of genic or multigenic thermolabile sex-determining systems, and could be combined with Biotechnology and Biomolecular Science, upstream thermal sensing and transduction to the epigenome to commit gonadal fate. UNSW Sydney, Sydney, NSW, Australia. Email: p.waters@unsw.edu.au Arthur Georges, Institute for Applied Ecology, University of Canberra, Canberra, Australia. Email: georges@aerg.canberra.edu.au Aurora Ruiz-Herrera, Departament de Biologia Cellular, Fisiologia i Immunologia, Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain. Email: aurora.ruizherrera@uab.cat INTRODUCTION ger conserved testis or ovary developmental pathways. The master switch that triggers male or female development is strikingly different Sex determination is the process by which a bipotential gonad is in different lineages, but gonad development is similar across verte- directed down a male (testis) or female (ovary) developmental path- brates and the complex molecular pathways of sexual differentiation way. In mammals, and many other vertebrates, sex is controlled by are relatively conserved.[1,2] sex chromosomes, defined by bearing sex-determining genes that trig- In all XY therian mammals, the Y-borne Sry is testis determining,[3] and in all ZW birds, the Z-borne Dmrt1 determines sex in a dosage- Abbreviations: 3C, chromosome conformation capture; 3D, three-dimensional; 3D-TSD, dependent manner.[4] However, reptiles present a truly impressive three dimensions of thermolabile sex determination; ESD, environmental sex determination; array of different sex-determining systems. Among reptiles with TSD, temperature-dependent sex determination; GSD, genetic sex determination; Hi-C, high-throughput chromosome conformation capture; TAD, topologically associated domain; genetic sex determination (GSD), turtles have male or female het- Pg, progesterone; PRC, polycomb repressive complex. erogamety (XY and ZW) (reviewed in Bista and Valenzuela[5] ), most Paul D. Waters and Aurora Ruiz-Herrera contributed equally to this study. snakes have female heterogamety (ZW or ZZW)[6] and both male and This is an open access article under the terms of the Creative Commons Attribution-NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes. © 2022 The Authors. BioEssays published by Wiley Periodicals LLC BioEssays. 2022;2200123. wileyonlinelibrary.com/journal/bies 1 of 8 https://doi.org/10.1002/bies.202200123 15211878, 0, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/bies.202200123 by Readcube (Labtiva Inc.), Wiley Online Library on [16/01/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License 2 of 8 WATERS ET AL . female heterogamety are observed in lizards (as well as XXY).[7] Sex order chromatin organization itself is thermosensitive, and changes chromosomes may be more or less differentiated. in 3D structure result in modulated expression of key sex genes In contrast to these systems of GSD, some vertebrates use envi- that impact gonadal fate. We have called this hypothesis the three ronmental triggers, such as temperature, to determine sex (environ- dimensions of thermolabile sex determination (3D-TSD for short). mental sex determination, ESD). The most common ESD mechanism, temperature-dependent sex determination (TSD), was discovered in Higher-order chromatin organization reptiles over 50 years ago,[8] challenging the orthodoxy of GSD that then prevailed. However, there are other systems of ESD involving Genomes are packaged into a chromatin structure, the regulation of different cues, including (but not exclusive to) social structure and which depends on different levels of organization, including (i) chemical resource availability (reviewed in Nagahama et al.[9] ). ESD raises major modifications of the DNA, (ii) modifications to the four core his- questions about how continuous variation in an external environmen- tones (H2A, H2B, H3 and H4) that comprise the nucleosomes around tal signal can shift cellular fate and commit development to either the which the DNA wraps and (iii) the 3D high-order organization of chro- ovary or testis molecular pathways. matin inside the nucleus that can change during the cell cycle and cell Decades of research have yielded limited insight to the mechanisms differentiation (Figure 1). by which temperature controls sex.[10,11] The problem of discovering Chromatin structure is maintained by DNA binding to histones, two epigenetic modification to an unknown differentially expressed gene each of H2A, H2B, H3 and H4, stabilised with H1. This structure is (or genes) seems intractable. It is not clear whether focus should be on modulated by the addition of different chemical groups to histone one candidate sex-determining gene, or a set of about 60 conserved tails (active or repressive marks), as well as by DNA methylation, sex genes (not necessarily on sex chromosomes in any species). Any and by their interactions with a host of other architectural factors, gene or gene product that promotes or shifts the trajectory of male enzymes, modifiers and transcription factors. For instance, dramatic and female development, even if it acts indirectly, is a candidate for change of chromatin conformation associated with histone modifica- thermal influence. There is also the possibility of a consensus or par- tion and DNA methylation mediates global transcriptional silencing of liamentary system, whereby the regulatory actions of many – perhaps the inactive X chromosome in mammals.[21] all – sex differentiation genes are collectively displaced by temper- The combination of high-resolution microscopy and chromosome ature to influence sexual outcomes.[12,13] Even if a common thread conformation capture (3C)-based methods (3C; 4C, 5C and Hi- was established by studies of traditional thermal sensing (e.g., cytosolic C) has revealed that the 3D chromatin structure is complex and Ca2+ and reactive oxygen species balance, alternative intron retention, dynamic. It includes chromosome territories in the interphase nucleus, etc.[11,14–16] ), this still leaves many candidate chromatin remodellers ‘open/active’ and ‘closed/inactive’ compartments (A and B), topologi- (epigenetic writers, readers or erasers) that can influence sex gene acti- cally associated domains (TADs), and looping interactions, which are vation or suppression through differential transcription and isoform established and maintained by structural proteins (i.e., cohesins and composition. Thus, identifying the mechanism of TSD has been, and CTCF)[20,22–25] (Figure 1). The compartmentalization of the genome remains, a difficult problem. in this manner partitions genomes into ‘regulatory neighbourhoods’ The fundamental questions underpinning TSD are (1) how is the by confining the activity of cis-regulatory elements to genes that fall thermal signal sensed? and (2) once captured, how is this signal within the same TAD.[26] It has been suggested that TAD bound- transduced into epigenetic change that releases or represses expres- aries can act as barriers between epigenetic states and that TADs sion of genes in sex-determining pathways? There have been several harbour-specific epigenetic signatures.[27] discoveries of changes in the expression, or transcripts, of down- Changes in the distribution of structural proteins and transcrip- stream genes, but little progress in identifying how the embryo senses tion factors, coupled with histone modifications associated with the temperature.[11,17] remodelling of high-order chromatin organization that impact on gene Recent research has shown that changes in the distribution of expression, occur during development and in the germline.[20,28–31] structural proteins coupled with epigenetic modifications (loading of This is highlighted by the knockout of epigenetic machinery (e.g., active/inactive marks) can have profound impact on gene expression histone deacetylases), which results in a changed epigenetic land- via a change to three-dimensional (3D) genome conformation. Indeed, scape that correlates with altered genomic contacts at promoters and such changes are an important part of cell differentiation during enhances that change gene expression.[32] In fact, many features of development.[18–20] 3D genome configuration of germ cells are highly dynamic, with cycli- What is not clear, however, is how directly or indirectly temperature cal transient chromatin–chromatin interactions that are established might affect the 3D structure of chromatin. For instance, temperature rapidly (reviewed in Refs. [24, 31]). The outcome is genomic plasticity has been suggested to have its effect indirectly via alterations in a that is poorly understood. temperature-sensitive ion channel that alters the balance of cytosolic Ca2+ and reactive oxygen species and affects epigenetic modifiers via phosphorylation of a control gene.[11] However, there remains the pos- Environmental 3D chromatin regulation sibility that increased temperature might alter 3D structures directly by disrupting existing promoter–enhancer interactions, or establishing Of the environmental stimuli that can influence chromatin regula- new ones. Here, we outline a novel hypothesis that proposes that high- tion, temperature is the most common. All organisms can respond 15211878, 0, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/bies.202200123 by Readcube (Labtiva Inc.), Wiley Online Library on [16/01/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License WATERS ET AL . 3 of 8 Loop Chromatin Cohesin TADs CTCF Nucleosomes Chromosome Territories B compartment B compartment A compartment F I G U R E 1 3D chromatin structure. The DNA wraps around histones forming nucleosomes, and constitutes the chromatin fibre. Chromatin fibres fold into chromatin loops forming topologically associated domains (TADs) with boundaries determined by cohesin complexes between CTCF convergent motifs. TADs are organised into A or B compartments, according to chromatin accessibility and transcriptional activity. Compartments are found within chromosome territories in nuclei. to temperature by activating a common transcriptional programme. tone modifications, promoted new long-range interactions that formed This heat shock response is well known to induce global changes new enhancer–promoter contacts that affected gene expression.[43] to gene regulation, as revealed in studies from human, mouse and Subsequent studies of human embryonic stem cells showed that Drosophila.[33–35] response to temperature resulted in changed enhancer–promoter Recently, the implications for direct 3D genome remodelling by interactions that correlated with a redistribution of RAD21 cohesin temperature have also been considered. New 3C-based methods that and CTCF.[44] Studies of hormone-induced changes show that spa- permit the study of how genomes remodel in response to the environ- tial structure of TADs plays a role in regulating the rapid transient ment at a fine scale (reviewed in Kumar et al.[36] ) have revealed that response to external signals. In vitro studies using T47D breast cancer plant genomes can remodel in response to salicylic acid[37] and proba- cells revealed that TAD border structures and their epigenetic modifi- bly light (reviewed in Perrella et al.[38] ). In mammals (e.g., mouse liver), cations can be rapidly modified (1 h) upon hormone (i.e., progesterone, TADs that harbour circadian genes switch between active and inac- Pg) stimulation, and this can occur over large genomic domains.[27] tive compartments at different times of the day, resulting in cycles of In this case, the Pg-activated receptor interacts with kinase signalling transcription modulation.[39] Thus, the response of 3D structure of the networks that regulate the expression of thousands of genes.[45] genome to environmental stimuli is nothing if not dynamic. Collectively, this evidence suggests that the thermoregulation of While our understanding of genome-level responses to environ- gene expression is tightly linked to chromatin remodelling, via changes mental changes is still limited, there is considerable variation in in structural proteins and transcription factors that can rapidly alter response to temperature across different organisms. Heat shock a host of interactions in response to environment (review in Kainth to cultured human and Drosophila cells caused dramatic tran- et al.[46] ). scriptional alteration without major changes in global chromatin architecture.[40] However, in plants and yeast, gene expression changes induced by heat stress were coupled with modification to 3D The three dimensions of sex genome structure.[37,41,42] In different Drosophila cells, heat stress induces a redistribution of architectural proteins that modulate TAD This relationship between thermoregulation and chromatin remod- boundaries.[43] This chromatin remodelling, coupled with covalent his- elling suggests that the genome senses the thermal signal via 15211878, 0, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/bies.202200123 by Readcube (Labtiva Inc.), Wiley Online Library on [16/01/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License 4 of 8 WATERS ET AL . temperature-induced chromatin remodelling, triggering either testis temperature disrupts DNA methylation of the Dmrt1 locus in ZW or ovary determination at early stages of development. embryos, resulting in the expression of this gene and pseudomale Here we propose that 3D genome conformation can respond development.[52] It is unknown how demethylation is mediated, but directly to temperature, resulting in chromatin remodelling in bipo- DNA methylation has profound effects on chromatin conformation[53] tential gonad precursor cells (Figure 2A). This could be reached by so its removal is likely to alter the 3D conformation and reactivate disrupting specific chromatin interactions, resulting in new genomic Dmrt1. contacts that change sex gene expression. The plethora of chromatin binding proteins, such as architectural (cohesins and CTCF), remod- ellers (epigenetic writers, readers and erasers) and transcription fac- Testing thermosensitive 3D conformation and sexual tors (review in Misteli[24] ), might be subjected to temperature-induced fate structural change. Altered folding of these proteins could promote changes in specific regulatory contacts. Significantly, recent studies in Examining the potential role of 3D genome remodelling in developing yeast have proposed that transcriptional condensates can rapidly and and/or sex-reversing embryonic gonads is now possible through the reversibly reconfigure the 3D genome in response to environmental implementation of an integrative approach that includes analysis of the conditions.[42] epigenome (histone modifications and DNA methylation) with genome Given the importance of the higher-order chromatin structure in structure (Hi-C) at different developmental stages and at different demarcating the limits of gene-regulatory domains, disturbances of temperatures, coupled with functional analysis (single cell RNA-seq) of this architecture would represent a means for rapid change in gene key sex genes. expression. Shifting TADs or compartment boundaries in response to For example, an excellent study system is brumation (akin to hiber- temperature would expose multiple genes to novel regulatory environ- nation) in the bearded dragon, during which thousands of genes are ments. This could break existing promotor enhancer contacts to turn differentially expressed compared to individuals at non-brumating genes off or establish new contacts to turn genes on (Figure 2A). Rel- temperatures.[54,55] Comparing genome structure in cold-brumating evant to our hypothesis is recent evidence of the role of chromatin individuals and warm individuals will reveal if the reptile genome remodelling during sex determination in mouse.[47] By integrating Hi-C has capacity to restructure in response to temperature. To determine and ChIP-seq data, the authors uncover rewiring of 3D enhancer hubs if the genome is remodelled in direct response to temperature, or during sex differentiation. In the light of this, we predict that the study whether change to the underlaying epigenetic code is responsible, of the structural and functional features that demarcate these dynamic profiling the epigenome (ChIP-seq/CUT&RUN/bisulfite sequencing), boundaries in different vertebrate lineages (i.e., reptiles with TSD) will including the detection of structural proteins such as cohesins and elucidate the mechanisms that govern higher-order genomic structure CTCF, in combination with Hi-C experiments in the developing rep- and function. tile gonad (at different temperatures) will be key. Both epigenome and This thermal sensing would induce genome remodelling in somatic Hi-C approaches could be conducted at different developmental time secretory gonadal cells (e.g., Sertoli, Leydig or granulosa) prior to sex points, from undifferentiated gonads through to developing ovaries differentiation, which would be maintained until the commitment of and testes after sexual fate is decided. This would reveal if the genome the gonad phenotype. Such thermosensitive chromatin interactions remodelled before the occurrence of changes in the epigenome or could bring key enhancers and promotors from distant locations into the distribution of structural proteins, which would indicate capac- close proximity (as recently proposed for transcriptional condensates ity for direct sensing of the thermal signal. Alternatively, if key in yeast[42] ) to alter gene expression in the gonad developmental path- genomic interactions were remodelled upon epigenetic change, this ways. Genes in newly formed compartments would then be directly would be indicative of an upstream sensing mechanism that results in regulated by this thermosensitive pathway. epigenome change that subsequently alters the high-order 3D genome Alternatively, the temperature may have a less direct effect on structure. chromatin structure. For instance, polycomb repressive complexes Research in reptile species with sex reversal might provide further (PRC) can alter chromatin structure (i.e., by H3K27me3 deposition) insights into the mechanisms involved. We propose that as well as resulting in changes of both cis and trans enhancer–promoter inter- acting in strictly TSD species, 3D TSD acts also in sex reversal sys- actions (reviewed in Illingworth[48] ), ultimately regulating potential tems when temperature overrides a genetic sex-determining gene. sex-determining genes[49,50] (Figure 2B). Therefore, non-canonical Particularly instructive might be two sex-reversing reptile species with isoforms of chromatin modifiers (i.e., ΔN-JARID2[15] ) with different opposite temperature-induced sex reversal. Pogona vitticeps has a ZZ affinity to PRC2 might act as a remodelling sensor, rather than a direct male:ZW female system in which ZZ develop as males at higher tem- regulator of sex genes. peratures. Bassiana duperreyi has the opposite system whereby XX Another indirect effect of temperature on chromatin might also individuals reverse to male at low temperatures.[56,57] In both cases, explain sex reversal in the half-smooth tongue sole. This fish has a we hypothesize that temperature acts directly to alter thermosensitive ZZ male:ZW female sex-determining system, in which a higher dosage 3D conformation, and affect the expression of influential genes in the of the Z-borne Dmrt1 directs male development.[51] However, higher sex differentiation pathway. 15211878, 0, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/bies.202200123 by Readcube (Labtiva Inc.), Wiley Online Library on [16/01/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License WATERS ET AL . 5 of 8 (A) 3D genome e rem remodelling modelling Directt sen nsing sensing TAD1 TAD2 TAD1 TAD2 TAD3 E1 P1 E2 P2 E3 P3 E1 P1 E2 P2 E3 P3 E: enhancers P: promoters Male Female On Off (B) Cytosol Phosphorylation Nucleus & relocalization 3D genome 1) TRPV STAT p STAT p structure channel STAT CaRe Ca2+ balance Ca2+ Ca2+ Calcium 3) Demethylase emeethylasse ions Ca2+ Splicing (act tivator)) (activator) KDM6B Ca2+ Ca2+ Ca2+ Cytosolic regulation ROS Promotor Prom motor Sex ge ene e gene Ca2+ + CLK4 PRC2 Methylase ? (repressor) JARID2 Sox9/AMH Wnt/Rspo1/ Foxl2 ΔN JARID2 Demethylase 2) Metabolism (activator) Male Female Phenotype F I G U R E 2 Sensing the thermal signal. (A) The genome could respond directly to temperature, resulting in sex genes being turned on or off. Three-dimensional genome (3D) remodelling could affect gene expression directly by disrupting establish (or establishing new) enhancer (E)–promoter (P) interactions within thermosensitive transcriptional condensates (pink circle). (B) A proposed route for how upstream thermal signals can be indirectly detected by the cell. Thermometer 1) TRPV channel activity is increased in response to temperature, increasing Ca2+ concentration in the cytosol. Thermometer 2) Higher temperature in reptiles increases metabolic rate and, therefore, ROS, which further activates TRPV channel activity. The thermosensitive CaRe balance results in phosphorylation (orange circles with p inside) of STAT, which subsequently relocates to the nucleus to affect epigenetic modifiers (e.g., KDM6B) and sex gene expression. Thermometer 3) Thermosensitive alternative splicing of epigenetic modifiers could directly alter the epigenetic landscapes that control to change sex gene expression. The thermo-sensitive CLK4 might regulate splicing. 15211878, 0, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/bies.202200123 by Readcube (Labtiva Inc.), Wiley Online Library on [16/01/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License 6 of 8 WATERS ET AL . CONCLUSION 4. Smith, C. A., Roeszler, K. N., Ohnesorg, T., Cummins, D. M., Farlie, P. G., Doran, T. J., & Sinclair, A. H. (2009). The avian Z-linked gene DMRT1 is required for male sex determination in the chicken. Nature, 461(7261), We propose that temperature can directly influence sex reversal by 267–271. https://doi.org/10.1038/nature08298 chromatin remodelling without invoking intermediate signal transduc- 5. Bista, B., & Valenzuela, N. (2020). Turtle insights into the evolu- tion. We hypothesize that the thermal signal could be directly sensed tion of the reptilian karyotype and the genomic architecture of by transcriptional condensates in the genome, resulting in altered sex determination. Genes (Basel), 11(4), 416. https://doi.org/10.3390/ genes11040416 enhancer–promoter interactions in the same TAD after compartment 6. Rovatsos, M., Augstenova, B., Altmanova, M., Sloboda, M., Kodym, switching. If a critical gene (or genes) in the sex-determining path- P., & Kratochvil, L. (2018). Triploid colubrid snake provides insight way switched compartments, the result could be to turn off (or on) into the mechanism of sex determination in advanced snakes. testis/ovary development, and ultimately reassign gonadal fate. This Sexual Development, 12(5), 251–255. https://doi.org/10.1159/ 000490124 fascinating possibility envisages a direct genomic thermal sensor that 7. Ezaz, T., Sarre, S. D., O’Meally, D., Graves, J. A. M., & Georges, A. skips intermediate signalling. (2009). Sex chromosome evolution in lizards: Independent origins and rapid transitions. Cytogenetic and Genome Research, 127(2-4), 249–260. AUTHOR CONTRIBUTIONS https://doi.org/10.1159/000300507 8. Charnier, M. (1966). Action of temperature on the sex ratio in the Aurora Ruiz-Herrera and Paul D. Waters wrote the first draft. Aurora Agama (Agamidae, Lacertilia) embryo. Comptes Rendus Des Seances De Ruiz-Herrera, Paul D. Waters, Arthur Georges and Jennifer A. Marshall La Societe De Biologie Et De Ses Filiales, 160(3), 620–622. Graves conceived the idea and commented on manuscript drafts. Sarah 9. Nagahama, Y., Chakraborty, T., Paul-Prasanth, B., Ohta, K., & L. Whiteley helped conceive ideas and prepared text for the supporting Nakamura, M. (2021). Sex determination, gonadal sex differentia- grant application, which we drew upon for this paper. Sarah L. Whiteley tion, and plasticity in vertebrate species. Physiological Reviews, 101(3), 1237–1308. https://doi.org/10.1152/physrev.00044.2019 also commented on manuscript drafts. 10. Weber, C., & Capel, B. (2021). Sex determination without sex chromo- somes. Philosophical Transactions of the Royal Society of London. Series ACKNOWLEDGEMENTS B: Biological Sciences, 376(1832), 20200109. https://doi.org/10.1098/ This research has received support from the Australian Research rstb.2020.0109 11. Castelli, M. A., Whiteley, S. L., Georges, A., & Holleley, C. E. (2020). Council [DP170101147 (A.G., P.D.W., J.A.M.G.), DP180100931 Cellular calcium and redox regulation: The mediator of vertebrate (P.D.W.), DP210103512 (P.D.W., J.A.M.G.), DP220101429 (A.G., P.D.W., environmental sex determination? Biological Reviews of the Cam- J.A.M.G., A.R.-H.)]. P.D.W. is supported by an NHMRC Project Grant bridge Philosophical Society, 95(3), 680–695. https://doi.org/10.1111/ (APP1156598). S.L.W. was funded by DP170101147 and CSIRO. A.R.- brv.12582 12. Crews, D., & Bull, J. J. (2009). Mode and tempo in environmental H. was supported by the Spanish Ministry of Science and Innovation sex determination in vertebrates. Seminars in Cell & Developmen- (PID2020-112557GB-I00). The authors are grateful to C. Vara for tal Biology, 20(3), 251–255. https://doi.org/10.1016/j.semcdb.2009. providing the first draft of Figure 1. 02.004 13. Georges, A., Ezaz, T., Quinn, A. E., & Sarre, S. D. (2010). Are reptiles predisposed to temperature-dependent sex determination? Sexual CONFLICT OF INTEREST Development, 4(1-2), 7–15. https://doi.org/10.1159/000279441 The authors declare no conflict of interest. 14. Haltenhof, T., Kotte, A., De Bortoli, F., Schiefer, S., Meinke, S., Emmerichs, A. K., Petermann, K. K., Timmermann, B., Imhof, P., Franz, DATA AVAILABILITY STATEMENT A., Loll, B., Wahl, M. C., Preußner, M., & Heyd, F. (2020). A conserved kinase-based body-temperature sensor globally controls alternative Data sharing is not applicable to this article as no datasets were splicing and gene expression. Molecular Cell, 78(1), 57–69.e4. https:// generated or analysed during the current study. doi.org/10.1016/j.molcel.2020.01.028 15. Deveson, I. W., Holleley, C. E., Blackburn, J., Graves, J. A. M., Mattick, ORCID J. S., Waters, P. D., & Georges, A. (2017). Differential intron reten- Paul D. Waters https://orcid.org/0000-0002-4689-8747 tion in Jumonji chromatin modifier genes is implicated in reptile temperature-dependent sex determination. Science Advances, 3(6), Aurora Ruiz-Herrera https://orcid.org/0000-0003-3868-6151 e1700731. https://doi.org/10.1126/sciadv.1700731 16. Whiteley, S. L., Wagner, S., Holleley, C. E., Deveson, I. W., Graves, J. 1. Capel, B. (2017). Vertebrate sex determination: Evolutionary plastic- A. M., & Georges, A. (2022). Truncated jarid2 and kdm6b transcripts ity of a fundamental switch. Nature Reviews Genetics, 18(11), 675–689. are associated with temperature-induced sex reversal during develop- https://doi.org/10.1038/nrg.2017.60 ment in a dragon lizard. Science Advances, 8(16), eabk0275. https://doi. 2. Herpin, A., & Schartl, M. (2015). Plasticity of gene-regulatory networks org/10.1126/sciadv.abk0275 controlling sex determination: Of masters, slaves, usual suspects, new- 17. Weber, C., Zhou, Y., Lee, J. G., Looger, L. L., Qian, G., Ge, C., & Capel, comers, and usurpators. EMBO Reports, 16(10), 1260–1274. https:// B. (2020). Temperature-dependent sex determination is mediated by doi.org/10.15252/embr.201540667 pSTAT3 repression of Kdm6b. Science, 368(6488), 303–306. https:// 3. Sinclair, A. H., Berta, P., Palmer, M. S., Hawkins, J. R., Griffiths, B. doi.org/10.1126/science.aaz4165 L., Smith, M. J., Foster, J. W., Frischauf, A.-M., Lovell-Badge, R., & 18. Ke, Y., Xu, Y., Chen, X., Feng, S., Liu, Z., Sun, Y., Yao, X., Li, F., Zhu, W., Goodfellow, P. N. (1990). A gene from the human sex-determining Gao, L., Chen, H., Du, Z., Xie, W., Xu, X., Huang, X., & Liu, J. (2017). 3D region encodes a protein with homology to a conserved DNA- chromatin structures of mature gametes and structural reprogram- binding motif. Nature, 346(6281), 240–244. https://doi.org/10.1038/ ming during mammalian embryogenesis. Cell, 170(2), 367–381.e20. 346240a0 https://doi.org/10.1016/j.cell.2017.06.029 15211878, 0, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/bies.202200123 by Readcube (Labtiva Inc.), Wiley Online Library on [16/01/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License WATERS ET AL . 7 of 8 19. Vallot, A., & Tachibana, K. (2020). The emergence of genome architec- nications, 11(1), 3419. https://doi.org/10.1038/s41467-020-17065- ture and zygotic genome activation. Current Opinion in Cell Biology, 64, z 50–57. https://doi.org/10.1016/j.ceb.2020.02.002 33. Mahat, D. B., Salamanca, H. H., Duarte, F. M., Danko, C. G., & Lis, J. T. 20. Vara, C., Paytuvi-Gallart, A., Cuartero, Y., Le Dily, F., Garcia, F., Salva- (2016). Mammalian heat shock response and mechanisms underlying Castro, J., Gómez-H, L., Julià, E., Moutinho, C., Aiese Cigliano, R., its genome-wide transcriptional regulation. Molecular Cell, 62(1), 63– Sanseverino, W., Fornas, O., Pendás, A. M., Heyn, H., Waters, P. D., 78. https://doi.org/10.1016/j.molcel.2016.02.025 Marti-Renom, M. A., & Ruiz-Herrera, A. (2019). Three-dimensional 34. Vihervaara, A., Sergelius, C., Vasara, J., Blom, M. A., Elsing, A. N., Roos- genomic structure and cohesin occupancy correlate with transcrip- Mattjus, P., & Sistonen, L. (2013). Transcriptional response to stress in tional activity during spermatogenesis. Cell Reports, 28(2), 352– the dynamic chromatin environment of cycling and mitotic cells. PNAS, 367.e9. https://doi.org/10.1016/j.celrep.2019.06.037 110(36), E3388–E3397. https://doi.org/10.1073/pnas.1305275110 21. Deng, X., Ma, W., Ramani, V., Hill, A., Yang, F., Ay, F., Berletch, J. B., Blau, 35. Yao, J., Munson, K. M., Webb, W. W., & Lis, J. T. (2006). Dynamics of heat C. A., Shendure, J., Duan, Z., Noble, W. S., & Disteche, C. M. (2015). shock factor association with native gene loci in living cells. Nature, Bipartite structure of the inactive mouse X chromosome. Genome 442(7106), 1050–1053. https://doi.org/10.1038/nature05025 Biology, 16, 152. https://doi.org/10.1186/s13059-015-0728-8 36. Kumar, S., Kaur, S., Seem, K., Kumar, S., & Mohapatra, T. (2021). Under- 22. Casa, V., Moronta Gines, M., Gade Gusmao, E., Slotman, J. A., Zirkel, standing 3D genome organization and its effect on transcriptional A., Josipovic, N., Oole, E., van IJcken, W. F. J., Houtsmuller, A. B., gene regulation under environmental stress in plant: A chromatin per- Papantonis, A., & Wendt, K. S. (2020). Redundant and specific roles spective. Frontiers in Cell and Developmental Biology, 9, 774719. https:// of cohesin STAG subunits in chromatin looping and transcriptional doi.org/10.3389/fcell.2021.774719 control. Genome Research, 30(4), 515–527. https://doi.org/10.1101/gr. 37. Yadav, V. K., Singh, S., Yadav, A., Agarwal, N., Singh, B., Jalmi, S. K., Yadav, 253211.119 V. K., Tiwari, V. K., Kumar, V., Singh, R., & Sawant, S. V. (2021). Stress 23. Dekker, J., Marti-Renom, M. A., & Mirny, L. A. (2013). Exploring the conditions modulate the chromatin interactions network in arabidop- three-dimensional organization of genomes: Interpreting chromatin sis. Frontiers in Genetics, 12, 799805. https://doi.org/10.3389/fgene. interaction data. Nature Reviews Genetics, 14(6), 390–403. https://doi. 2021.799805 org/10.1038/nrg3454 38. Perrella, G., Zioutopoulou, A., Headland, L. R., & Kaiserli, E. (2020). The 24. Misteli, T. (2020). The self-organizing genome: Principles of genome impact of light and temperature on chromatin organization and plant architecture and function. Cell, 183(1), 28–45. https://doi.org/10. adaptation. Journal of Experimental Botany, 71(17), 5247–5255. https:// 1016/j.cell.2020.09.014 doi.org/10.1093/jxb/eraa154 25. Rao, S. S., Huntley, M. H., Durand, N. C., Stamenova, E. K., Bochkov, I. 39. Furlan-Magaril, M., Ando-Kuri, M., Arzate-Mejia, R. G., Morf, J., Cairns, D., Robinson, J. T., Sanborn, A. L., Machol, I., Omer, A. D., Lander, E. S., & J., Roman-Figueroa, A., Tenorio-Hernández, L., Poot-Hernández, A. C., Aiden, E. L. (2014). A 3D map of the human genome at kilobase resolu- Andrews, S., Várnai, C., Virk, B., Wingett, S. W., & Fraser, P. (2021). tion reveals principles of chromatin looping. Cell, 159(7), 1665–1680. The global and promoter-centric 3D genome organization temporally https://doi.org/10.1016/j.cell.2014.11.021 resolved during a circadian cycle. Genome Biology, 22(1), 162. https:// 26. Long, H. K., Prescott, S. L., & Wysocka, J. (2016). Ever-changing doi.org/10.1186/s13059-021-02374-3 landscapes: Transcriptional enhancers in development and evolu- 40. Ray, J., Munn, P. R., Vihervaara, A., Lewis, J. J., Ozer, A., Danko, C. tion. Cell, 167(5), 1170–1187. https://doi.org/10.1016/j.cell.2016. G., & Lis, J. T. (2019). Chromatin conformation remains stable upon 09.018 extensive transcriptional changes driven by heat shock. PNAS, 116(39), 27. Le Dily, F., Bau, D., Pohl, A., Vicent, G. P., Serra, F., Soronellas, D., 19431–19439. https://doi.org/10.1073/pnas.1901244116 Castellano, G., Wright, R. H. G., Ballare, C., Filion, G., Marti-Renom, 41. Chowdhary, S., Kainth, A. S., & Gross, D. S. (2017). Heat shock protein M. A., & Beato, M. (2014). Distinct structural transitions of chromatin genes undergo dynamic alteration in their three-dimensional structure topological domains correlate with coordinated hormone-induced and genome organization in response to thermal stress. Molecular and gene regulation. Genes & Development, 28(19), 2151–2162. https://doi. Cellular Biology, 37(24). https://doi.org/10.1128/MCB.00292-17 org/10.1101/gad.241422.114 42. Chowdhary, S., Kainth, A. S., Paracha, S., Gross, D. S., & Pincus, 28. Alavattam, K. G., Maezawa, S., Sakashita, A., Khoury, H., Barski, A., D. (2022). Inducible transcriptional condensates drive 3D genome Kaplan, N., & Namekawa, S. H. (2019). Attenuated chromatin com- reorganization in the heat shock response. Molecular Cell, 82(22), partmentalization in meiosis and its maturation in sperm development. 4386–4399.e7. https://doi.org/10.1016/j.molcel.2022.10.013 Nature Structural & Molecular Biology, 26(3), 175–184. https://doi.org/ 43. Li, L., Lyu, X., Hou, C., Takenaka, N., Nguyen, H. Q., Ong, C. T., Cubeñas- 10.1038/s41594-019-0189-y Potts, C., Hu, M., Lei, E. P., Bosco, G., Qin, Z. S., & Corces, V. G. 29. Patel, L., Kang, R., Rosenberg, S. C., Qiu, Y., Raviram, R., Chee, S., Hu, (2015). Widespread rearrangement of 3D chromatin organization R., Ren, B., Cole, F., & Corbett, K. D. (2019). Dynamic reorganization of underlies polycomb-mediated stress-induced silencing. Molecular Cell, the genome shapes the recombination landscape in meiotic prophase. 58(2), 216–231. https://doi.org/10.1016/j.molcel.2015.02.023 Nature Structural & Molecular Biology, 26(3), 164–174. https://doi.org/ 44. Lyu, X., Rowley, M. J., & Corces, V. G. (2018). Architectural proteins 10.1038/s41594-019-0187-0 and pluripotency factors cooperate to orchestrate the transcrip- 30. Vara, C., Paytuvi-Gallart, A., Cuartero, Y., Alvarez-Gonzalez, L., Marin- tional response of hESCs to temperature stress. Molecular Cell, 71(6), Gual, L., Garcia, F., Florit-Sabater, B., Capilla, L., Sanchéz-Guillén, R. A., 940–955.e7. https://doi.org/10.1016/j.molcel.2018.07.012 Sarrate, Z., Aiese Cigliano, R., Sanseverino, W., Searle, J. B., Ventura, J., 45. Wright, R. H., Castellano, G., Bonet, J., Le Dily, F., Font-Mateu, J., Marti-Renom, M. A., Le Dily, F., & Ruiz-Herrera, A. (2021). The impact Ballare, C., Nacht, A. S., Soronellas, D., Oliva, B., & Beato, M. (2012). of chromosomal fusions on 3D genome folding and recombination in CDK2-dependent activation of PARP-1 is required for hormonal the germ line. Nature Communications, 12(1), 2981. https://doi.org/10. gene regulation in breast cancer cells. Genes & Development, 26(17), 1038/s41467-021-23270-1 1972–1983. https://doi.org/10.1101/gad.193193.112 31. Vara, C., & Ruiz-Herrera, A. (2022). Unpacking chromatin remodelling 46. Kainth, A. S., Chowdhary, S., Pincus, D., & Gross, D. S. (2021). Primor- in germ cells: Implications for development and evolution. Trends in dial super-enhancers: Heat shock-induced chromatin organization in Genetics, 38(5), 422–425. https://doi.org/10.1016/j.tig.2021.10.007 yeast. Trends in Cell Biology, 31(10), 801–813. https://doi.org/10.1016/ 32. Goodman, J. V., Yamada, T., Yang, Y., Kong, L., Wu, D. Y., Zhao, G., Gabel, j.tcb.2021.04.004 H. W., & Bonni, A. (2020). The chromatin remodeling enzyme Chd4 47. Mota-Gómez, I., Rodríguez, J. A., Dupont, S., Lao, O., Jedamzick, regulates genome architecture in the mouse brain. Nature Commu- J., Kuhn, R., Lacadie, S., García-Moreno, S. A., Hurtado, A., Acemel, 15211878, 0, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/bies.202200123 by Readcube (Labtiva Inc.), Wiley Online Library on [16/01/2023]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License 8 of 8 WATERS ET AL . R. D., Capel, B., Marti-Renom, M. A., & Lupiáñez, D. G. (2022). in Structural Biology, 75, 102430. https://doi.org/10.1016/j.sbi.2022. Sex-determining 3D regulatory hubs revealed by genome spatial 102430 auto-correlation analysis. bioRxiv. https://doi.org/10.1101/2022.11. 54. Capraro, A., O’Meally, D., Waters, S. A., Patel, H. R., Georges, A., & 18.516861 Waters, P. D. (2019). Waking the sleeping dragon: Gene expression 48. Illingworth, R. S. (2019). Chromatin folding and nuclear architecture: profiling reveals adaptive strategies of the hibernating reptile Pogona PRC1 function in 3D. Current Opinion in Genetics & Development, 55, vitticeps. BMC Genomics [Electronic Resource], 20(1), 460. https://doi. 82–90. https://doi.org/10.1016/j.gde.2019.06.006 org/10.1186/s12864-019-5750-x 49. Ge, C., Ye, J., Weber, C., Sun, W., Zhang, H., Zhou, Y., Cai, C., Qian, 55. Capraro, A., O’Meally, D., Waters, S. A., Patel, H. R., Georges, A., & G., & Capel, B. (2018). The histone demethylase KDM6B regulates Waters, P. D. (2020). MicroRNA dynamics during hibernation of the temperature-dependent sex determination in a turtle species. Science, Australian central bearded dragon (Pogona vitticeps). Scientific Reports, 360(6389), 645–648. https://doi.org/10.1126/science.aap8328 10(1), 17854. https://doi.org/10.1038/s41598-020-73706-9 50. Ge, C., Ye, J., Zhang, H., Zhang, Y., Sun, W., Sang, Y., Capel, B., & Qian, 56. Dissanayake, D. S. B., Holleley, C. E., & Georges, A. (2021). Effects of G. (2017). Dmrt1 induces the male pathway in a turtle species with natural nest temperatures on sex reversal and sex ratios in an Aus- temperature-dependent sex determination. Development (Cambridge, tralian alpine skink. Scientific Reports, 11(1), 20093. https://doi.org/10. England), 144(12), 2222–2233. https://doi.org/10.1242/dev.152033 1038/s41598-021-99702-1 51. Chen, S., Zhang, G., Shao, C., Huang, Q., Liu, G., Zhang, P., Song, W., An, 57. Radder, R. S., Quinn, A. E., Georges, A., Sarre, S. D., & Shine, R. (2008). N., Chalopin, D., Volff, J.-N., Hong, Y., Li, Q., Sha, Z., Zhou, H., Xie, M., Genetic evidence for co-occurrence of chromosomal and thermal sex- Yu, Q., Liu, Y., Xiang, H., Wang, N., . . . Wang, J. (2014). Whole-genome determining systems in a lizard. Biology Letters, 4(2), 176–178. https:// sequence of a flatfish provides insights into ZW sex chromosome evo- doi.org/10.1098/rsbl.2007.0583 lution and adaptation to a benthic lifestyle. Nature Genetics, 46(3), 253–260. https://doi.org/10.1038/ng.2890 52. Shao, C., Li, Q., Chen, S., Zhang, P., Lian, J., Hu, Q., Sun, B., Jin, L., Liu, S., Wang, Z., Zhao, H., Jin, Z., Liang, Z., Li, Y., Zheng, Q., Zhang, Y., Wang, J., How to cite this article: Waters, P. D., Graves, J. A. M., & Zhang, G. (2014). Epigenetic modification and inheritance in sexual Whiteley, S. L., Georges, A., & Ruiz-Herrera, A. (2022). Three reversal of fish. Genome Research, 24(4), 604–615. https://doi.org/10. 1101/gr.162172.113 Dimensions Of Thermolabile Sex Determination. BioEssays, 53. Li, S., Peng, Y., & Panchenko, A. R. (2022). DNA methylation: Pre- e2200123. https://doi.org/10.1002/bies.202200123 cise modulation of chromatin structure and dynamics. Current Opinion