Pub Date : 2024-06-18DOI: 10.1038/s41594-024-01337-z
Xiaozhan Qu, Shuo Zhao, Chanjuan Wan, Lei Zhu, Tuo Ji, Paolo Rossi, Junfeng Wang, Charalampos G. Kalodimos, Chao Wang, Weiya Xu, Chengdong Huang
Molecular chaperone heat shock protein 90 (Hsp90) is a ubiquitous regulator that fine-tunes and remodels diverse client proteins, exerting profound effects on normal biology and diseases. Unraveling the mechanistic details of Hsp90’s function requires atomic-level insights into its client interactions throughout the adenosine triphosphate-coupled functional cycle. However, the structural details of the initial encounter complex in the chaperone cycle, wherein Hsp90 adopts an open conformation while engaging with the client, remain elusive. Here, using nuclear magnetic resonance spectroscopy, we determined the solution structure of Hsp90 in its open state, bound to a disordered client. Our findings reveal that Hsp90 uses two distinct binding sites, collaborating synergistically to capture discrete hydrophobic segments within client proteins. This bipartite interaction generates a versatile complex that facilitates rapid conformational sampling. Moreover, our investigations spanning various clients and Hsp90 orthologs demonstrate a pervasive mechanism used by Hsp90 orthologs to accommodate the vast array of client proteins. Collectively, our work contributes to establish a unified conceptual and mechanistic framework, elucidating the intricate interplay between Hsp90 and its clients.
{"title":"Structural basis for the dynamic chaperoning of disordered clients by Hsp90","authors":"Xiaozhan Qu, Shuo Zhao, Chanjuan Wan, Lei Zhu, Tuo Ji, Paolo Rossi, Junfeng Wang, Charalampos G. Kalodimos, Chao Wang, Weiya Xu, Chengdong Huang","doi":"10.1038/s41594-024-01337-z","DOIUrl":"https://doi.org/10.1038/s41594-024-01337-z","url":null,"abstract":"<p>Molecular chaperone heat shock protein 90 (Hsp90) is a ubiquitous regulator that fine-tunes and remodels diverse client proteins, exerting profound effects on normal biology and diseases. Unraveling the mechanistic details of Hsp90’s function requires atomic-level insights into its client interactions throughout the adenosine triphosphate-coupled functional cycle. However, the structural details of the initial encounter complex in the chaperone cycle, wherein Hsp90 adopts an open conformation while engaging with the client, remain elusive. Here, using nuclear magnetic resonance spectroscopy, we determined the solution structure of Hsp90 in its open state, bound to a disordered client. Our findings reveal that Hsp90 uses two distinct binding sites, collaborating synergistically to capture discrete hydrophobic segments within client proteins. This bipartite interaction generates a versatile complex that facilitates rapid conformational sampling. Moreover, our investigations spanning various clients and Hsp90 orthologs demonstrate a pervasive mechanism used by Hsp90 orthologs to accommodate the vast array of client proteins. Collectively, our work contributes to establish a unified conceptual and mechanistic framework, elucidating the intricate interplay between Hsp90 and its clients.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-06-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141334417","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-13DOI: 10.1038/s41594-024-01332-4
Feng Wang, Qing He, Nina Y. Yao, Michael E. O’Donnell, Huilin Li
Humans have three different proliferating cell nuclear antigen (PCNA) clamp-loading complexes: RFC and CTF18-RFC load PCNA onto DNA, but ATAD5-RFC can only unload PCNA from DNA. The underlying structural basis of ATAD5-RFC unloading is unknown. We show here that ATAD5 has two unique locking loops that appear to tie the complex into a rigid structure, and together with a domain that plugs the DNA-binding chamber, prevent conformation changes required for DNA binding, likely explaining why ATAD5-RFC is exclusively a PCNA unloader. These features are conserved in the yeast PCNA unloader Elg1-RFC. We observe intermediates in which PCNA bound to ATAD5-RFC exists as a closed planar ring, a cracked spiral or a gapped spiral. Surprisingly, ATAD5-RFC can open a PCNA gap between PCNA protomers 2 and 3, different from the PCNA protomers 1 and 3 gap observed in all previously characterized clamp loaders.
{"title":"The human ATAD5 has evolved unique structural elements to function exclusively as a PCNA unloader","authors":"Feng Wang, Qing He, Nina Y. Yao, Michael E. O’Donnell, Huilin Li","doi":"10.1038/s41594-024-01332-4","DOIUrl":"https://doi.org/10.1038/s41594-024-01332-4","url":null,"abstract":"<p>Humans have three different proliferating cell nuclear antigen (PCNA) clamp-loading complexes: RFC and CTF18-RFC load PCNA onto DNA, but ATAD5-RFC can only unload PCNA from DNA. The underlying structural basis of ATAD5-RFC unloading is unknown. We show here that ATAD5 has two unique locking loops that appear to tie the complex into a rigid structure, and together with a domain that plugs the DNA-binding chamber, prevent conformation changes required for DNA binding, likely explaining why ATAD5-RFC is exclusively a PCNA unloader. These features are conserved in the yeast PCNA unloader Elg1-RFC. We observe intermediates in which PCNA bound to ATAD5-RFC exists as a closed planar ring, a cracked spiral or a gapped spiral. Surprisingly, ATAD5-RFC can open a PCNA gap between PCNA protomers 2 and 3, different from the PCNA protomers 1 and 3 gap observed in all previously characterized clamp loaders.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-06-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141315573","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-12DOI: 10.1038/s41594-024-01334-2
Hossein Batebi, Guillermo Pérez-Hernández, Sabrina N. Rahman, Baoliang Lan, Antje Kamprad, Mingyu Shi, David Speck, Johanna K. S. Tiemann, Ramon Guixà-González, Franziska Reinhardt, Peter F. Stadler, Makaía M. Papasergi-Scott, Georgios Skiniotis, Patrick Scheerer, Brian K. Kobilka, Jesper M. Mathiesen, Xiangyu Liu, Peter W. Hildebrand
G-protein-coupled receptors (GPCRs) activate heterotrimeric G proteins by promoting guanine nucleotide exchange. Here, we investigate the coupling of G proteins with GPCRs and describe the events that ultimately lead to the ejection of GDP from its binding pocket in the Gα subunit, the rate-limiting step during G-protein activation. Using molecular dynamics simulations, we investigate the temporal progression of structural rearrangements of GDP-bound Gs protein (Gs·GDP; hereafter GsGDP) upon coupling to the β2-adrenergic receptor (β2AR) in atomic detail. The binding of GsGDP to the β2AR is followed by long-range allosteric effects that significantly reduce the energy needed for GDP release: the opening of α1-αF helices, the displacement of the αG helix and the opening of the α-helical domain. Signal propagation to the Gs occurs through an extended receptor interface, including a lysine-rich motif at the intracellular end of a kinked transmembrane helix 6, which was confirmed by site-directed mutagenesis and functional assays. From this β2AR–GsGDP intermediate, Gs undergoes an in-plane rotation along the receptor axis to approach the β2AR–Gsempty state. The simulations shed light on how the structural elements at the receptor–G-protein interface may interact to transmit the signal over 30 Å to the nucleotide-binding site. Our analysis extends the current limited view of nucleotide-free snapshots to include additional states and structural features responsible for signaling and G-protein coupling specificity.
G 蛋白偶联受体(GPCR)通过促进鸟嘌呤核苷酸交换来激活异三聚体 G 蛋白。在这里,我们研究了 G 蛋白与 GPCR 的耦合,并描述了最终导致 GDP 从 Gα 亚基的结合袋中排出的事件,这是 G 蛋白激活过程中的限速步骤。通过分子动力学模拟,我们研究了与 GDP 结合的 Gs 蛋白(Gs-GDP,以下简称 GsGDP)在与 β2-肾上腺素能受体(β2AR)耦合时结构重排的时间进展细节。GsGDP 与 β2AR 结合后会产生长程异构效应,大大降低 GDP 释放所需的能量:α1-αF 螺旋的打开、αG 螺旋的移位和 α-helical 结构域的打开。信号通过一个扩展的受体界面传播到 Gs,该界面包括位于扭结跨膜螺旋 6 胞内端的一个富含赖氨酸的基团,这一点已通过定点突变和功能测试得到证实。从这个 β2AR-GsGDP 中间体开始,Gs 沿着受体轴进行平面内旋转,以接近 β2AR-Gsempty 状态。模拟揭示了受体-蛋白质界面上的结构元素如何相互作用,将信号传输 30 Å 以上至核苷酸结合位点。我们的分析扩展了目前无核苷酸快照的有限视角,纳入了更多的状态和结构特征,这些状态和特征负责信号传导和 G 蛋白偶联特异性。
{"title":"Mechanistic insights into G-protein coupling with an agonist-bound G-protein-coupled receptor","authors":"Hossein Batebi, Guillermo Pérez-Hernández, Sabrina N. Rahman, Baoliang Lan, Antje Kamprad, Mingyu Shi, David Speck, Johanna K. S. Tiemann, Ramon Guixà-González, Franziska Reinhardt, Peter F. Stadler, Makaía M. Papasergi-Scott, Georgios Skiniotis, Patrick Scheerer, Brian K. Kobilka, Jesper M. Mathiesen, Xiangyu Liu, Peter W. Hildebrand","doi":"10.1038/s41594-024-01334-2","DOIUrl":"https://doi.org/10.1038/s41594-024-01334-2","url":null,"abstract":"<p>G-protein-coupled receptors (GPCRs) activate heterotrimeric G proteins by promoting guanine nucleotide exchange. Here, we investigate the coupling of G proteins with GPCRs and describe the events that ultimately lead to the ejection of GDP from its binding pocket in the Gα subunit, the rate-limiting step during G-protein activation. Using molecular dynamics simulations, we investigate the temporal progression of structural rearrangements of GDP-bound G<sub>s</sub> protein (G<sub>s</sub>·GDP; hereafter G<sub>s</sub><sup>GDP</sup>) upon coupling to the β<sub>2</sub>-adrenergic receptor (β<sub>2</sub>AR) in atomic detail. The binding of G<sub>s</sub><sup>GDP</sup> to the β<sub>2</sub>AR is followed by long-range allosteric effects that significantly reduce the energy needed for GDP release: the opening of α1-αF helices, the displacement of the αG helix and the opening of the α-helical domain. Signal propagation to the G<sub>s</sub> occurs through an extended receptor interface, including a lysine-rich motif at the intracellular end of a kinked transmembrane helix 6, which was confirmed by site-directed mutagenesis and functional assays. From this β<sub>2</sub>AR–G<sub>s</sub><sup>GDP</sup> intermediate, G<sub>s</sub> undergoes an in-plane rotation along the receptor axis to approach the β<sub>2</sub>AR–G<sub>s</sub><sup>empty</sup> state. The simulations shed light on how the structural elements at the receptor–G-protein interface may interact to transmit the signal over 30 Å to the nucleotide-binding site. Our analysis extends the current limited view of nucleotide-free snapshots to include additional states and structural features responsible for signaling and G-protein coupling specificity.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-06-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141309030","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-04DOI: 10.1038/s41594-024-01320-8
Vivien Horváth, Raquel Garza, Marie E. Jönsson, Pia A. Johansson, Anita Adami, Georgia Christoforidou, Ofelia Karlsson, Laura Castilla Vallmanya, Symela Koutounidou, Patricia Gerdes, Ninoslav Pandiloski, Christopher H. Douse, Johan Jakobsson
SVA (SINE (short interspersed nuclear element)–VNTR (variable number of tandem repeats)–Alu) retrotransposons remain active in humans and contribute to individual genetic variation. Polymorphic SVA alleles harbor gene regulatory potential and can cause genetic disease. However, how SVA insertions are controlled and functionally impact human disease is unknown. Here we dissect the epigenetic regulation and influence of SVAs in cellular models of X-linked dystonia parkinsonism (XDP), a neurodegenerative disorder caused by an SVA insertion at the TAF1 locus. We demonstrate that the KRAB zinc finger protein ZNF91 establishes H3K9me3 and DNA methylation over SVAs, including polymorphic alleles, in human neural progenitor cells. The resulting mini-heterochromatin domains attenuate the cis-regulatory impact of SVAs. This is critical for XDP pathology; removal of local heterochromatin severely aggravates the XDP molecular phenotype, resulting in increased TAF1 intron retention and reduced expression. Our results provide unique mechanistic insights into how human polymorphic transposon insertions are recognized and how their regulatory impact is constrained by an innate epigenetic defense system.
SVA(SINE(短穿插核元素)-VNTR(可变串联重复序列)-Alu)反转座子在人类中依然活跃,并导致个体遗传变异。多态的 SVA 等位基因具有基因调控潜能,可导致遗传疾病。然而,SVA插入如何受控并对人类疾病产生功能性影响尚不清楚。在这里,我们对 X 连锁肌张力障碍性帕金森病(XDP)细胞模型中 SVA 的表观遗传调控和影响进行了剖析,这是一种由 TAF1 基因座上的 SVA 插入引起的神经退行性疾病。我们证明了 KRAB 锌指蛋白 ZNF91 能在人类神经祖细胞中的 SVA(包括多态等位基因)上建立 H3K9me3 和 DNA 甲基化。由此产生的微型异染色质域减弱了 SVA 的顺式调控影响。这对 XDP 病理学至关重要;去除局部异染色质会严重加剧 XDP 分子表型,导致 TAF1 内含子保留增加和表达减少。我们的研究结果为人类多态转座子插入如何被识别以及它们的调控影响如何被先天表观遗传防御系统所限制提供了独特的机理见解。
{"title":"Mini-heterochromatin domains constrain the cis-regulatory impact of SVA transposons in human brain development and disease","authors":"Vivien Horváth, Raquel Garza, Marie E. Jönsson, Pia A. Johansson, Anita Adami, Georgia Christoforidou, Ofelia Karlsson, Laura Castilla Vallmanya, Symela Koutounidou, Patricia Gerdes, Ninoslav Pandiloski, Christopher H. Douse, Johan Jakobsson","doi":"10.1038/s41594-024-01320-8","DOIUrl":"https://doi.org/10.1038/s41594-024-01320-8","url":null,"abstract":"<p>SVA (SINE (short interspersed nuclear element)–VNTR (variable number of tandem repeats)–<i>Alu</i>) retrotransposons remain active in humans and contribute to individual genetic variation. Polymorphic SVA alleles harbor gene regulatory potential and can cause genetic disease. However, how SVA insertions are controlled and functionally impact human disease is unknown. Here we dissect the epigenetic regulation and influence of SVAs in cellular models of X-linked dystonia parkinsonism (XDP), a neurodegenerative disorder caused by an SVA insertion at the <i>TAF1</i> locus. We demonstrate that the KRAB zinc finger protein ZNF91 establishes H3K9me3 and DNA methylation over SVAs, including polymorphic alleles, in human neural progenitor cells. The resulting mini-heterochromatin domains attenuate the <i>cis</i>-regulatory impact of SVAs. This is critical for XDP pathology; removal of local heterochromatin severely aggravates the XDP molecular phenotype, resulting in increased <i>TAF1</i> intron retention and reduced expression. Our results provide unique mechanistic insights into how human polymorphic transposon insertions are recognized and how their regulatory impact is constrained by an innate epigenetic defense system.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141246269","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-04DOI: 10.1038/s41594-024-01328-0
W. Dylan Hale, Alejandra Montaño Romero, Cuauhtemoc U. Gonzalez, Vasanthi Jayaraman, Albert Y. Lau, Richard L. Huganir, Edward C. Twomey
Excitatory neurotransmission is principally mediated by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-subtype ionotropic glutamate receptors (AMPARs). Negative allosteric modulators are therapeutic candidates that inhibit AMPAR activation and can compete with positive modulators to control AMPAR function through unresolved mechanisms. Here we show that allosteric inhibition pushes AMPARs into a distinct state that prevents both activation and positive allosteric modulation. We used cryo-electron microscopy to capture AMPARs bound to glutamate, while a negative allosteric modulator, GYKI-52466, and positive allosteric modulator, cyclothiazide, compete for control of the AMPARs. GYKI-52466 binds in the ion channel collar and inhibits AMPARs by decoupling the ligand-binding domains from the ion channel. The rearrangement of the ligand-binding domains ruptures the cyclothiazide site, preventing positive modulation. Our data provide a framework for understanding allostery of AMPARs and for rational design of therapeutics targeting AMPARs in neurological diseases.
{"title":"Allosteric competition and inhibition in AMPA receptors","authors":"W. Dylan Hale, Alejandra Montaño Romero, Cuauhtemoc U. Gonzalez, Vasanthi Jayaraman, Albert Y. Lau, Richard L. Huganir, Edward C. Twomey","doi":"10.1038/s41594-024-01328-0","DOIUrl":"https://doi.org/10.1038/s41594-024-01328-0","url":null,"abstract":"<p>Excitatory neurotransmission is principally mediated by α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-subtype ionotropic glutamate receptors (AMPARs). Negative allosteric modulators are therapeutic candidates that inhibit AMPAR activation and can compete with positive modulators to control AMPAR function through unresolved mechanisms. Here we show that allosteric inhibition pushes AMPARs into a distinct state that prevents both activation and positive allosteric modulation. We used cryo-electron microscopy to capture AMPARs bound to glutamate, while a negative allosteric modulator, GYKI-52466, and positive allosteric modulator, cyclothiazide, compete for control of the AMPARs. GYKI-52466 binds in the ion channel collar and inhibits AMPARs by decoupling the ligand-binding domains from the ion channel. The rearrangement of the ligand-binding domains ruptures the cyclothiazide site, preventing positive modulation. Our data provide a framework for understanding allostery of AMPARs and for rational design of therapeutics targeting AMPARs in neurological diseases.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141246303","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-06-04DOI: 10.1038/s41594-024-01325-3
Charbel Alfeghaly, Gaël Castel, Emmanuel Cazottes, Madeleine Moscatelli, Eva Moinard, Miguel Casanova, Juliette Boni, Kasturi Mahadik, Jenna Lammers, Thomas Freour, Louis Chauviere, Carla Piqueras, Ruben Boers, Joachim Boers, Joost Gribnau, Laurent David, Jean-François Ouimette, Claire Rougeulle
XIST (X-inactive specific transcript) long noncoding RNA (lncRNA) is responsible for X chromosome inactivation (XCI) in placental mammals, yet it accumulates on both X chromosomes in human female preimplantation embryos without triggering X chromosome silencing. The XACT (X-active coating transcript) lncRNA coaccumulates with XIST on active X chromosomes and may antagonize XIST function. Here, we used human embryonic stem cells in a naive state of pluripotency to assess the function of XIST and XACT in shaping the X chromosome chromatin and transcriptional landscapes during preimplantation development. We show that XIST triggers the deposition of polycomb-mediated repressive histone modifications and dampens the transcription of most X-linked genes in a SPEN-dependent manner, while XACT deficiency does not significantly affect XIST activity or X-linked gene expression. Our study demonstrates that XIST is functional before XCI, confirms the existence of a transient process of X chromosome dosage compensation and reveals that XCI and dampening rely on the same set of factors.
XIST(X-inactive specific transcript)长非编码 RNA(lncRNA)在胎盘哺乳动物中负责 X 染色体失活(XCI),但它会在人类女性植入前胚胎的两条 X 染色体上积累,而不会引发 X 染色体沉默。XACT(X-活性包被转录本)lncRNA与XIST共同聚集在活性X染色体上,可能会拮抗XIST的功能。在这里,我们利用处于幼稚多能状态的人类胚胎干细胞,评估了XIST和XACT在植入前发育过程中塑造X染色体染色质和转录景观的功能。我们的研究表明,XIST 会触发多聚酶介导的抑制性组蛋白修饰沉积,并以一种依赖 SPEN 的方式抑制大多数 X 连锁基因的转录,而 XACT 的缺乏并不会显著影响 XIST 的活性或 X 连锁基因的表达。我们的研究表明,XIST 在 XCI 之前就已发挥作用,证实了 X 染色体剂量补偿过程的存在,并揭示了 XCI 和抑制作用依赖于同一组因子。
{"title":"XIST dampens X chromosome activity in a SPEN-dependent manner during early human development","authors":"Charbel Alfeghaly, Gaël Castel, Emmanuel Cazottes, Madeleine Moscatelli, Eva Moinard, Miguel Casanova, Juliette Boni, Kasturi Mahadik, Jenna Lammers, Thomas Freour, Louis Chauviere, Carla Piqueras, Ruben Boers, Joachim Boers, Joost Gribnau, Laurent David, Jean-François Ouimette, Claire Rougeulle","doi":"10.1038/s41594-024-01325-3","DOIUrl":"https://doi.org/10.1038/s41594-024-01325-3","url":null,"abstract":"<p>XIST (X-inactive specific transcript) long noncoding RNA (lncRNA) is responsible for X chromosome inactivation (XCI) in placental mammals, yet it accumulates on both X chromosomes in human female preimplantation embryos without triggering X chromosome silencing. The XACT (X-active coating transcript) lncRNA coaccumulates with XIST on active X chromosomes and may antagonize XIST function. Here, we used human embryonic stem cells in a naive state of pluripotency to assess the function of XIST and XACT in shaping the X chromosome chromatin and transcriptional landscapes during preimplantation development. We show that XIST triggers the deposition of polycomb-mediated repressive histone modifications and dampens the transcription of most X-linked genes in a SPEN-dependent manner, while XACT deficiency does not significantly affect XIST activity or X-linked gene expression. Our study demonstrates that XIST is functional before XCI, confirms the existence of a transient process of X chromosome dosage compensation and reveals that XCI and dampening rely on the same set of factors.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141246284","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-05-29DOI: 10.1038/s41594-024-01322-6
Junjiao Yang, Chan-I Chung, Jessica Koach, Hongjiang Liu, Ambuja Navalkar, Hao He, Zhimin Ma, Qian Zhao, Xiaoyu Yang, Liang He, Tanja Mittag, Yin Shen, William A. Weiss, Xiaokun Shu
Dysregulation and enhanced expression of MYC transcription factors (TFs) including MYC and MYCN contribute to the majority of human cancers. For example, MYCN is amplified up to several hundredfold in high-risk neuroblastoma. The resulting overexpression of N-myc aberrantly activates genes that are not activated at low N-myc levels and drives cell proliferation. Whether increasing N-myc levels simply mediates binding to lower-affinity binding sites in the genome or fundamentally changes the activation process remains unclear. One such activation mechanism that could become important above threshold levels of N-myc is the formation of aberrant transcriptional condensates through phase separation. Phase separation has recently been linked to transcriptional regulation, but the extent to which it contributes to gene activation remains an open question. Here we characterized the phase behavior of N-myc and showed that it can form dynamic condensates that have transcriptional hallmarks. We tested the role of phase separation in N-myc-regulated transcription by using a chemogenetic tool that allowed us to compare non-phase-separated and phase-separated conditions at equivalent N-myc levels, both of which showed a strong impact on gene expression compared to no N-myc expression. Interestingly, we discovered that only a small percentage (<3%) of N-myc-regulated genes is further modulated by phase separation but that these events include the activation of key oncogenes and the repression of tumor suppressors. Indeed, phase separation increases cell proliferation, corroborating the biological effects of the transcriptional changes. However, our results also show that >97% of N-myc-regulated genes are not affected by N-myc phase separation, demonstrating that soluble complexes of TFs with the transcriptional machinery are sufficient to activate transcription.
{"title":"MYC phase separation selectively modulates the transcriptome","authors":"Junjiao Yang, Chan-I Chung, Jessica Koach, Hongjiang Liu, Ambuja Navalkar, Hao He, Zhimin Ma, Qian Zhao, Xiaoyu Yang, Liang He, Tanja Mittag, Yin Shen, William A. Weiss, Xiaokun Shu","doi":"10.1038/s41594-024-01322-6","DOIUrl":"https://doi.org/10.1038/s41594-024-01322-6","url":null,"abstract":"<p>Dysregulation and enhanced expression of <i>MYC</i> transcription factors (TFs) including <i>MYC</i> and <i>MYCN</i> contribute to the majority of human cancers. For example, <i>MYCN</i> is amplified up to several hundredfold in high-risk neuroblastoma. The resulting overexpression of N-myc aberrantly activates genes that are not activated at low N-myc levels and drives cell proliferation. Whether increasing N-myc levels simply mediates binding to lower-affinity binding sites in the genome or fundamentally changes the activation process remains unclear. One such activation mechanism that could become important above threshold levels of N-myc is the formation of aberrant transcriptional condensates through phase separation. Phase separation has recently been linked to transcriptional regulation, but the extent to which it contributes to gene activation remains an open question. Here we characterized the phase behavior of N-myc and showed that it can form dynamic condensates that have transcriptional hallmarks. We tested the role of phase separation in N-myc-regulated transcription by using a chemogenetic tool that allowed us to compare non-phase-separated and phase-separated conditions at equivalent N-myc levels, both of which showed a strong impact on gene expression compared to no N-myc expression. Interestingly, we discovered that only a small percentage (<3%) of N-myc-regulated genes is further modulated by phase separation but that these events include the activation of key oncogenes and the repression of tumor suppressors. Indeed, phase separation increases cell proliferation, corroborating the biological effects of the transcriptional changes. However, our results also show that >97% of N-myc-regulated genes are not affected by N-myc phase separation, demonstrating that soluble complexes of TFs with the transcriptional machinery are sufficient to activate transcription.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-05-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141165114","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-05-28DOI: 10.1038/s41594-024-01323-5
Jochen Kuper, Tamsanqa Hove, Sarah Maidl, Hermann Neitz, Florian Sauer, Maximilian Kempf, Till Schroeder, Elke Greiter, Claudia Höbartner, Caroline Kisker
The superfamily 2 helicase XPD is a central component of the general transcription factor II H (TFIIH), which is essential for transcription and nucleotide excision DNA repair (NER). Within these two processes, the helicase function of XPD is vital for NER but not for transcription initiation, where XPD acts only as a scaffold for other factors. Using cryo-EM, we deciphered one of the most enigmatic steps in XPD helicase action: the active separation of double-stranded DNA (dsDNA) and its stalling upon approaching a DNA interstrand cross-link, a highly toxic form of DNA damage. The structure shows how dsDNA is separated and reveals a highly unusual involvement of the Arch domain in active dsDNA separation. Combined with mutagenesis and biochemical analyses, we identified distinct functional regions important for helicase activity. Surprisingly, those areas also affect core TFIIH translocase activity, revealing a yet unencountered function of XPD within the TFIIH scaffold. In summary, our data provide a universal basis for NER bubble formation, XPD damage verification and XPG incision.
超家族 2 螺旋酶 XPD 是一般转录因子 II H(TFIIH)的核心成分,它对转录和核苷酸切割 DNA 修复(NER)至关重要。在这两个过程中,XPD 的螺旋酶功能对 NER 至关重要,但对转录起始并不重要,在转录起始过程中,XPD 只充当其他因子的支架。我们利用低温电子显微镜破译了 XPD 螺旋酶作用中最神秘的步骤之一:主动分离双链 DNA(dsDNA),并在接近 DNA 链间交联(一种剧毒的 DNA 损伤形式)时停滞。该结构显示了dsDNA是如何分离的,并揭示了Arch结构域在主动分离dsDNA过程中极不寻常的参与。结合诱变和生化分析,我们确定了对螺旋酶活性非常重要的不同功能区。令人惊讶的是,这些区域也影响了 TFIIH 核心转位酶的活性,揭示了 XPD 在 TFIIH 支架中尚未遇到的功能。总之,我们的数据为 NER 气泡形成、XPD 损伤验证和 XPG 切割提供了一个普遍基础。
{"title":"XPD stalled on cross-linked DNA provides insight into damage verification","authors":"Jochen Kuper, Tamsanqa Hove, Sarah Maidl, Hermann Neitz, Florian Sauer, Maximilian Kempf, Till Schroeder, Elke Greiter, Claudia Höbartner, Caroline Kisker","doi":"10.1038/s41594-024-01323-5","DOIUrl":"https://doi.org/10.1038/s41594-024-01323-5","url":null,"abstract":"<p>The superfamily 2 helicase XPD is a central component of the general transcription factor II H (TFIIH), which is essential for transcription and nucleotide excision DNA repair (NER). Within these two processes, the helicase function of XPD is vital for NER but not for transcription initiation, where XPD acts only as a scaffold for other factors. Using cryo-EM, we deciphered one of the most enigmatic steps in XPD helicase action: the active separation of double-stranded DNA (dsDNA) and its stalling upon approaching a DNA interstrand cross-link, a highly toxic form of DNA damage. The structure shows how dsDNA is separated and reveals a highly unusual involvement of the Arch domain in active dsDNA separation. Combined with mutagenesis and biochemical analyses, we identified distinct functional regions important for helicase activity. Surprisingly, those areas also affect core TFIIH translocase activity, revealing a yet unencountered function of XPD within the TFIIH scaffold. In summary, our data provide a universal basis for NER bubble formation, XPD damage verification and XPG incision.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-05-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141159521","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-05-24DOI: 10.1038/s41594-024-01314-6
Yingxia Hu, Krista A. Delviks-Frankenberry, Chunxiang Wu, Fidel Arizaga, Vinay K. Pathak, Yong Xiong
HIV-1 Vif recruits host cullin-RING-E3 ubiquitin ligase and CBFβ to degrade the cellular APOBEC3 antiviral proteins through diverse interactions. Recent evidence has shown that Vif also degrades the regulatory subunits PPP2R5(A–E) of cellular protein phosphatase 2A to induce G2/M cell cycle arrest. As PPP2R5 proteins bear no functional or structural resemblance to A3s, it is unclear how Vif can recognize different sets of proteins. Here we report the cryogenic-electron microscopy structure of PPP2R5A in complex with HIV-1 Vif–CBFβ–elongin B–elongin C at 3.58 Å resolution. The structure shows PPP2R5A binds across the Vif molecule, with biochemical and cellular studies confirming a distinct Vif–PPP2R5A interface that partially overlaps with those for A3s. Vif also blocks a canonical PPP2R5A substrate-binding site, indicating that it suppresses the phosphatase activities through both degradation-dependent and degradation-independent mechanisms. Our work identifies critical Vif motifs regulating the recognition of diverse A3 and PPP2R5A substrates, whereby disruption of these host–virus protein interactions could serve as potential targets for HIV-1 therapeutics.
{"title":"Structural insights into PPP2R5A degradation by HIV-1 Vif","authors":"Yingxia Hu, Krista A. Delviks-Frankenberry, Chunxiang Wu, Fidel Arizaga, Vinay K. Pathak, Yong Xiong","doi":"10.1038/s41594-024-01314-6","DOIUrl":"https://doi.org/10.1038/s41594-024-01314-6","url":null,"abstract":"<p>HIV-1 Vif recruits host cullin-RING-E3 ubiquitin ligase and CBFβ to degrade the cellular APOBEC3 antiviral proteins through diverse interactions. Recent evidence has shown that Vif also degrades the regulatory subunits PPP2R5(A–E) of cellular protein phosphatase 2A to induce G2/M cell cycle arrest. As PPP2R5 proteins bear no functional or structural resemblance to A3s, it is unclear how Vif can recognize different sets of proteins. Here we report the cryogenic-electron microscopy structure of PPP2R5A in complex with HIV-1 Vif–CBFβ–elongin B–elongin C at 3.58 Å resolution. The structure shows PPP2R5A binds across the Vif molecule, with biochemical and cellular studies confirming a distinct Vif–PPP2R5A interface that partially overlaps with those for A3s. Vif also blocks a canonical PPP2R5A substrate-binding site, indicating that it suppresses the phosphatase activities through both degradation-dependent and degradation-independent mechanisms. Our work identifies critical Vif motifs regulating the recognition of diverse A3 and PPP2R5A substrates, whereby disruption of these host–virus protein interactions could serve as potential targets for HIV-1 therapeutics.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-05-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141092183","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-05-23DOI: 10.1038/s41594-024-01313-7
Maximilian Stötzel, Chieh-Yu Cheng, Ibrahim A. IIik, Abhishek Sampath Kumar, Persia Akbari Omgba, Vera A. van der Weijden, Yufei Zhang, Martin Vingron, Alexander Meissner, Tuğçe Aktaş, Helene Kretzmer, Aydan Bulut-Karslioğlu
Dormancy is an essential biological process for the propagation of many life forms through generations and stressful conditions. Early embryos of many mammals are preservable for weeks to months within the uterus in a dormant state called diapause, which can be induced in vitro through mTOR inhibition. Cellular strategies that safeguard original cell identity within the silent genomic landscape of dormancy are not known. Here we show that the protection of cis-regulatory elements from silencing is key to maintaining pluripotency in the dormant state. We reveal a TET–transcription factor axis, in which TET-mediated DNA demethylation and recruitment of methylation-sensitive transcription factor TFE3 drive transcriptionally inert chromatin adaptations during dormancy transition. Perturbation of TET activity compromises pluripotency and survival of mouse embryos under dormancy, whereas its enhancement improves survival rates. Our results reveal an essential mechanism for propagating the cellular identity of dormant cells, with implications for regeneration and disease.
{"title":"TET activity safeguards pluripotency throughout embryonic dormancy","authors":"Maximilian Stötzel, Chieh-Yu Cheng, Ibrahim A. IIik, Abhishek Sampath Kumar, Persia Akbari Omgba, Vera A. van der Weijden, Yufei Zhang, Martin Vingron, Alexander Meissner, Tuğçe Aktaş, Helene Kretzmer, Aydan Bulut-Karslioğlu","doi":"10.1038/s41594-024-01313-7","DOIUrl":"https://doi.org/10.1038/s41594-024-01313-7","url":null,"abstract":"<p>Dormancy is an essential biological process for the propagation of many life forms through generations and stressful conditions. Early embryos of many mammals are preservable for weeks to months within the uterus in a dormant state called diapause, which can be induced in vitro through mTOR inhibition. Cellular strategies that safeguard original cell identity within the silent genomic landscape of dormancy are not known. Here we show that the protection of <i>cis</i>-regulatory elements from silencing is key to maintaining pluripotency in the dormant state. We reveal a TET–transcription factor axis, in which TET-mediated DNA demethylation and recruitment of methylation-sensitive transcription factor TFE3 drive transcriptionally inert chromatin adaptations during dormancy transition. Perturbation of TET activity compromises pluripotency and survival of mouse embryos under dormancy, whereas its enhancement improves survival rates. Our results reveal an essential mechanism for propagating the cellular identity of dormant cells, with implications for regeneration and disease.</p>","PeriodicalId":18822,"journal":{"name":"Nature structural & molecular biology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-05-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141085341","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}