f68c45b20190e3435226500149c74b9d pmc_34209946.pdf 2a7e47e05aa9c5446c44482d151d105fe1da37ef pmc_34209946.pdf 556bbd11b0c13323e858c67708295b377a114187264476c73aa97190b6543ad2 pmc_34209946.pdf Title: Synonymous Codon Pair Recoding of the HIV-1 env Gene Affects Virus Replication Capacity Subject: Synonymous codon pair deoptimization is an efficient strategy for virus attenuation; however, the underlying mechanism remains controversial. Here, we optimized and deoptimized the codon pair bias (CPB) of the human immunodeficiency virus type 1 (HIV-1) envelope (env) gene to investigate the influence of env synonymous CPB recoding on virus replication capacity, as well as the potential mechanism. We found that env CPB deoptimization did not always generate attenuation, whereas CPB optimization attenuated virus replication in MT-4 cells. Furthermore, virus attenuation correlated with reduced Env protein production but not with decreased viral RNA synthesis. Remarkably, in our model, increasing the number of CpG dinucleotides in the 5' end of env did not reduce the replication capacity of HIV-1. These results indicate that factors other than CPB or CpG content may have impacted the viral fitness of the synonymously recoded study variants. Our findings provide evidence that CPB recoding-associated attenuation can affect translation efficiency. Moreover, we demonstrated that an increased number of CpGs in the 5' end of HIV-1 env is not always associated with reduced virus replication capacity. Keywords: codon pair bias; synonymous gene recoding; HIV-1; envelope; fitness; attenuation Author: Ana Jordan-Paiz, Sandra Franco and Miguel Angel Martinez Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Thu Jul 1 05:00:40 2021 CEST ModDate: Thu Jul 1 05:04:00 2021 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 10 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 972374 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- PMZFBW+VnURWPalladioL Type 1 Custom yes yes no 10 0 GKXOSR+URWPalladioL-Ital Type 1 Custom yes yes no 15 0 XVNIUD+URWPalladioL-Bold Type 1 Custom yes yes no 20 0 YOUFHF+URWPalladioL-BoldItal Type 1 Custom yes yes no 24 0 TAJDLB+URWPalladioL-Roma Type 1 Custom yes yes no 28 0 IFEDSM+CMSY10 Type 1 Builtin yes yes no 32 0 PFKOHU+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 40 0 MCBCCI+SourceSansRoman-Regular Type 1C Custom yes yes no 62 0 MCBCCM+SourceSansRoman.613wght Type 1C Custom yes yes no 66 0 MCBCCO+SourceSansRoman-Bold Type 1C Custom yes yes no 70 0 OVNEXW+EURM10 Type 1 Builtin yes yes no 85 0 Jhove (Rel. 1.22.1, 2019-04-17) Date: 2023-10-21 02:37:05 CEST RepresentationInformation: pmc_34209946.pdf ReportingModule: PDF-hul, Rel. 1.12.1 (2019-04-17) LastModified: 2023-03-31 12:14:24 CEST Size: 972374 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 279 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@262b2c86 Item: Title: Materials and Methods Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@371a67ec Children: Item: Title: Cell Lines Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@5ed828d Item: Title: Generation of Synthetic Infectious HIV-1 Env Variants Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@50d0686 Item: Title: Replication Capacity Assays Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7a3d45bd Item: Title: Plasmids and HIV-1 Env Expression Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1e7c7811 Item: Title: Immunoblot Analyses and Antibodies Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77ec78b9 Item: Title: Real-Time Quantitative PCR Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1a3869f4 Item: Title: Statistical Analysis Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@a38d7a3 Item: Title: Results Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77f99a05 Item: Title: Discussion Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@63440df3 Item: Title: References Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@3aeaafa6 Info: Title: Synonymous Codon Pair Recoding of the HIV-1 env Gene Affects Virus Replication Capacity Author: Ana Jordan-Paiz, Sandra Franco and Miguel Angel Martinez Subject: Synonymous codon pair deoptimization is an efficient strategy for virus attenuation; however, the underlying mechanism remains controversial. Here, we optimized and deoptimized the codon pair bias (CPB) of the human immunodeficiency virus type 1 (HIV-1) envelope (env) gene to investigate the influence of env synonymous CPB recoding on virus replication capacity, as well as the potential mechanism. We found that env CPB deoptimization did not always generate attenuation, whereas CPB optimization attenuated virus replication in MT-4 cells. Furthermore, virus attenuation correlated with reduced Env protein production but not with decreased viral RNA synthesis. Remarkably, in our model, increasing the number of CpG dinucleotides in the 5' end of env did not reduce the replication capacity of HIV-1. These results indicate that factors other than CPB or CpG content may have impacted the viral fitness of the synonymously recoded study variants. Our findings provide evidence that CPB recoding-associated attenuation can affect translation efficiency. Moreover, we demonstrated that an increased number of CpGs in the 5' end of HIV-1 env is not always associated with reduced virus replication capacity. Keywords: codon pair bias; synonymous gene recoding; HIV-1; envelope; fitness; attenuation Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Thu Jul 01 05:00:40 CEST 2021 ModDate: Thu Jul 01 05:04:00 CEST 2021 ID: 0xf20b1a93dbf1c3a4fbb79216f6d761ae, 0xf20b1a93dbf1c3a4fbb79216f6d761ae Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: IFEDSM+CMSY10 FontSubset: true FirstChar: 0 LastChar: 48 FontDescriptor: FontName: IFEDSM+CMSY10 Flags: Symbolic FontBBox: -29, -960, 1116, 775 FontFile: true Font: BaseFont: XVNIUD+URWPalladioL-Bold FontSubset: true FirstChar: 3 LastChar: 134 FontDescriptor: FontName: XVNIUD+URWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: OVNEXW+EURM10 FontSubset: true FirstChar: 22 LastChar: 22 FontDescriptor: FontName: OVNEXW+EURM10 Flags: Symbolic FontBBox: -32, -243, 1060, 720 FontFile: true Font: BaseFont: YOUFHF+URWPalladioL-BoldItal FontSubset: true FirstChar: 101 LastChar: 118 FontDescriptor: FontName: YOUFHF+URWPalladioL-BoldItal Flags: Symbolic FontBBox: -170, -300, 1073, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: PMZFBW+VnURWPalladioL FontSubset: true FirstChar: 161 LastChar: 226 FontDescriptor: FontName: PMZFBW+VnURWPalladioL Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: TAJDLB+URWPalladioL-Roma FontSubset: true FirstChar: 2 LastChar: 252 FontDescriptor: FontName: TAJDLB+URWPalladioL-Roma Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: GKXOSR+URWPalladioL-Ital FontSubset: true FirstChar: 45 LastChar: 121 FontDescriptor: FontName: GKXOSR+URWPalladioL-Ital Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true Pages: Page: Sequence: 1 Annotations: Annotation: Subtype: Link Rect: 35, 759, 122, 795 Annotation: Subtype: Link Rect: 515, 759, 560, 795 Annotation: Subtype: Link Rect: 188, 659, 199, 673 Annotation: Subtype: Link Rect: 334, 659, 345, 673 Annotation: Subtype: Link Rect: 98, 358, 154, 367 Annotation: Subtype: Link Rect: 35, 346, 131, 355 Annotation: Subtype: Link Rect: 35, 162, 97, 184 Annotation: Subtype: Link Rect: 153, 89, 154, 98 Annotation: Subtype: Link Rect: 35, 77, 147, 86 Annotation: Subtype: Link Rect: 516, 319, 523, 328 ActionDest: 113 Annotation: Subtype: Link Rect: 523, 319, 530, 328 ActionDest: 113 Annotation: Subtype: Link Rect: 216, 231, 223, 240 ActionDest: 113 Annotation: Subtype: Link Rect: 226, 231, 238, 240 ActionDest: 113 Annotation: Subtype: Link Rect: 426, 193, 433, 202 ActionDest: 113 Annotation: Subtype: Link Rect: 434, 193, 441, 202 ActionDest: 113 Annotation: Subtype: Link Rect: 525, 155, 537, 164 ActionDest: 113 Annotation: Subtype: Link Rect: 441, 143, 453, 152 ActionDest: 113 Annotation: Subtype: Link Rect: 378, 118, 390, 127 ActionDest: 113 Annotation: Subtype: Link Rect: 476, 118, 488, 127 ActionDest: 113 Annotation: Subtype: Link Rect: 526, 118, 538, 127 ActionDest: 113 Annotation: Subtype: Link Rect: 300, 105, 312, 114 ActionDest: 116 Annotation: Subtype: Link Rect: 399, 105, 411, 114 ActionDest: 116 Annotation: Subtype: Link Rect: 418, 80, 430, 89 ActionDest: 116 Annotation: Subtype: Link Rect: 431, 80, 443, 89 ActionDest: 116 Annotation: Subtype: Link Rect: 104, 17, 244, 28 Annotation: Subtype: Link Rect: 419, 17, 558, 28 Annotation: Subtype: Link Rect: 35, 423, 87, 442 Page: Sequence: 2 Annotations: Annotation: Subtype: Link Rect: 491, 711, 498, 721 ActionDest: 113 Annotation: Subtype: Link Rect: 324, 686, 336, 695 ActionDest: 113 Annotation: Subtype: Link Rect: 412, 649, 424, 658 ActionDest: 113 Annotation: Subtype: Link Rect: 548, 611, 555, 620 ActionDest: 113 Annotation: Subtype: Link Rect: 303, 586, 310, 595 ActionDest: 113 Annotation: Subtype: Link Rect: 311, 586, 318, 595 ActionDest: 113 Annotation: Subtype: Link Rect: 477, 306, 558, 318 Annotation: Subtype: Link Rect: 536, 245, 543, 255 ActionDest: 113 Annotation: Subtype: Link Rect: 544, 245, 556, 255 ActionDest: 116 Page: Sequence: 3 Annotations: Annotation: Subtype: Link Rect: 461, 722, 468, 731 ActionDest: 113 Annotation: Subtype: Link Rect: 534, 684, 541, 693 ActionDest: 113 Annotation: Subtype: Link Rect: 544, 684, 556, 693 ActionDest: 113 Annotation: Subtype: Link Rect: 468, 596, 475, 605 ActionDest: 113 Annotation: Subtype: Link Rect: 478, 596, 490, 605 ActionDest: 113 Annotation: Subtype: Link Rect: 216, 533, 223, 542 ActionDest: 113 Annotation: Subtype: Link Rect: 484, 496, 496, 505 ActionDest: 116 Annotation: Subtype: Link Rect: 447, 382, 454, 391 ActionDest: 113 Annotation: Subtype: Link Rect: 407, 194, 414, 203 ActionDest: 113 Page: Sequence: 4 Annotations: Annotation: Subtype: Link Rect: 483, 584, 495, 593 ActionDest: 113 Annotation: Subtype: Link Rect: 320, 518, 327, 530 ActionDest: 89 Annotation: Subtype: Link Rect: 240, 493, 247, 505 ActionDest: 89 Annotation: Subtype: Link Rect: 286, 443, 293, 455 ActionDest: 89 Annotation: Subtype: Link Rect: 282, 142, 289, 151 ActionDest: 113 Annotation: Subtype: Link Rect: 490, 129, 502, 138 ActionDest: 113 Annotation: Subtype: Link Rect: 517, 114, 524, 126 ActionDest: 89 Annotation: Subtype: Link Rect: 191, 76, 198, 88 ActionDest: 89 Page: Sequence: 5 Annotations: Annotation: Subtype: Link Rect: 360, 732, 367, 744 ActionDest: 89 Annotation: Subtype: Link Rect: 338, 707, 345, 719 ActionDest: 89 Annotation: Subtype: Link Rect: 362, 644, 369, 656 ActionDest: 92 Annotation: Subtype: Link Rect: 363, 314, 370, 323 ActionDest: 113 Annotation: Subtype: Link Rect: 211, 289, 218, 298 ActionDest: 113 Annotation: Subtype: Link Rect: 549, 224, 556, 236 ActionDest: 89 Annotation: Subtype: Link Rect: 403, 148, 410, 161 ActionDest: 98 Annotation: Subtype: Link Rect: 484, 86, 491, 98 ActionDest: 89 Annotation: Subtype: Link Rect: 478, 60, 485, 73 ActionDest: 98 Page: Sequence: 6 Annotations: Annotation: Subtype: Link Rect: 498, 732, 505, 745 ActionDest: 98 Annotation: Subtype: Link Rect: 353, 707, 360, 720 ActionDest: 98 Annotation: Subtype: Link Rect: 397, 660, 404, 669 ActionDest: 113 Annotation: Subtype: Link Rect: 371, 123, 378, 136 ActionDest: 104 Annotation: Subtype: Link Rect: 319, 73, 326, 85 ActionDest: 104 Page: Sequence: 7 Annotations: Annotation: Subtype: Link Rect: 529, 419, 536, 428 ActionDest: 113 Annotation: Subtype: Link Rect: 537, 419, 543, 428 ActionDest: 113 Annotation: Subtype: Link Rect: 544, 419, 555, 428 ActionDest: 113 Annotation: Subtype: Link Rect: 246, 406, 253, 415 ActionDest: 113 Annotation: Subtype: Link Rect: 253, 406, 260, 415 ActionDest: 113 Annotation: Subtype: Link Rect: 261, 406, 273, 415 ActionDest: 113 Annotation: Subtype: Link Rect: 195, 381, 202, 390 ActionDest: 113 Annotation: Subtype: Link Rect: 203, 381, 215, 390 ActionDest: 113 Annotation: Subtype: Link Rect: 204, 343, 215, 352 ActionDest: 116 Annotation: Subtype: Link Rect: 216, 343, 228, 352 ActionDest: 116 Annotation: Subtype: Link Rect: 364, 318, 376, 327 ActionDest: 116 Annotation: Subtype: Link Rect: 544, 218, 556, 227 ActionDest: 116 Annotation: Subtype: Link Rect: 197, 105, 209, 114 ActionDest: 116 Annotation: Subtype: Link Rect: 390, 92, 397, 101 ActionDest: 113 Annotation: Subtype: Link Rect: 518, 80, 530, 89 ActionDest: 113 Page: Sequence: 8 Annotations: Annotation: Subtype: Link Rect: 482, 735, 489, 744 ActionDest: 113 Annotation: Subtype: Link Rect: 490, 735, 502, 744 ActionDest: 113 Annotation: Subtype: Link Rect: 502, 735, 514, 744 ActionDest: 116 Annotation: Subtype: Link Rect: 515, 735, 527, 744 ActionDest: 116 Annotation: Subtype: Link Rect: 392, 672, 399, 681 ActionDest: 113 Annotation: Subtype: Link Rect: 400, 672, 407, 681 ActionDest: 113 Annotation: Subtype: Link Rect: 370, 647, 377, 656 ActionDest: 113 Annotation: Subtype: Link Rect: 289, 634, 296, 643 ActionDest: 113 Annotation: Subtype: Link Rect: 240, 534, 247, 543 ActionDest: 113 Annotation: Subtype: Link Rect: 544, 496, 556, 505 ActionDest: 113 Annotation: Subtype: Link Rect: 235, 471, 247, 480 ActionDest: 113 Annotation: Subtype: Link Rect: 221, 383, 233, 392 ActionDest: 116 Annotation: Subtype: Link Rect: 310, 358, 317, 367 ActionDest: 113 Annotation: Subtype: Link Rect: 321, 321, 333, 330 ActionDest: 116 Annotation: Subtype: Link Rect: 333, 320, 345, 330 ActionDest: 116 Annotation: Subtype: Link Rect: 202, 308, 209, 317 ActionDest: 113 Annotation: Subtype: Link Rect: 390, 195, 397, 204 ActionDest: 113 Annotation: Subtype: Link Rect: 397, 195, 404, 204 ActionDest: 113 Annotation: Subtype: Link Rect: 405, 195, 417, 204 ActionDest: 113 Page: Sequence: 9 Annotations: Annotation: Subtype: Link Rect: 419, 696, 560, 707 Annotation: Subtype: Link Rect: 166, 684, 260, 695 Annotation: Subtype: Link Rect: 329, 453, 367, 464 Annotation: Subtype: Link Rect: 373, 453, 409, 464 Annotation: Subtype: Link Rect: 197, 431, 235, 441 Annotation: Subtype: Link Rect: 241, 431, 277, 441 Annotation: Subtype: Link Rect: 519, 407, 557, 418 Annotation: Subtype: Link Rect: 59, 396, 95, 406 Annotation: Subtype: Link Rect: 424, 372, 462, 383 Annotation: Subtype: Link Rect: 181, 338, 218, 349 Annotation: Subtype: Link Rect: 127, 315, 164, 326 Annotation: Subtype: Link Rect: 188, 280, 225, 291 Annotation: Subtype: Link Rect: 59, 246, 97, 257 Annotation: Subtype: Link Rect: 103, 246, 139, 257 Annotation: Subtype: Link Rect: 412, 223, 450, 234 Annotation: Subtype: Link Rect: 380, 200, 418, 211 Annotation: Subtype: Link Rect: 361, 177, 399, 188 Annotation: Subtype: Link Rect: 405, 177, 441, 188 Annotation: Subtype: Link Rect: 477, 153, 514, 165 Annotation: Subtype: Link Rect: 104, 131, 142, 141 Annotation: Subtype: Link Rect: 240, 107, 277, 118 Annotation: Subtype: Link Rect: 283, 107, 319, 118 Annotation: Subtype: Link Rect: 394, 84, 431, 95 Annotation: Subtype: Link Rect: 437, 84, 473, 95 Page: Sequence: 10 Annotations: Annotation: Subtype: Link Rect: 131, 710, 168, 720 Annotation: Subtype: Link Rect: 104, 676, 142, 686 Annotation: Subtype: Link Rect: 148, 676, 184, 686 Annotation: Subtype: Link Rect: 379, 652, 417, 663 Annotation: Subtype: Link Rect: 519, 640, 557, 651 Annotation: Subtype: Link Rect: 59, 630, 95, 640 Annotation: Subtype: Link Rect: 231, 594, 269, 605 Annotation: Subtype: Link Rect: 275, 594, 311, 605 Annotation: Subtype: Link Rect: 519, 571, 557, 582 Annotation: Subtype: Link Rect: 59, 561, 95, 571 Annotation: Subtype: Link Rect: 474, 537, 512, 548 Annotation: Subtype: Link Rect: 518, 537, 554, 548 Annotation: Subtype: Link Rect: 233, 502, 271, 513 Annotation: Subtype: Link Rect: 277, 502, 313, 513 Annotation: Subtype: Link Rect: 336, 491, 373, 502 Annotation: Subtype: Link Rect: 460, 468, 498, 479 Annotation: Subtype: Link Rect: 160, 445, 197, 456 Annotation: Subtype: Link Rect: 179, 411, 217, 421 Annotation: Subtype: Link Rect: 450, 387, 487, 398 Checksum: a60f7e7e Type: CRC32 Checksum: f68c45b20190e3435226500149c74b9d Type: MD5 Checksum: 2a7e47e05aa9c5446c44482d151d105fe1da37ef Type: SHA-1