Biochimica et Biophysica Acta-Gene Regulatory Mechanisms最新文献_第3页

Growth rate is related to elongation of RNA polymerase II transcription in Saccharomyces cerevisiae 酿酒酵母菌的生长速率与RNA聚合酶II转录的伸长有关。

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-06-08 DOI: 10.1016/j.bbagrm.2025.195100

A.I. Garrido-Godino , R. González , M. Martín-Expósito , S. Chávez , J.E. Pérez-Ortín , F. Navarro

Cells must adapt to changing environmental conditions to maintain their fitness and to compete with other genotypes during the natural selection process. The growth rate (GR) is a determining factor in this competition, and it influences gene expression. Some genes increase mRNA levels, while others decrease with the GR. mRNA levels depend on the dynamic balance between their synthesis by RNA polymerase II and their degradation rates. RNA polymerase I and III are also influenced by the GR because they transcribe protein synthesis machinery required to make proteins that increase cell mass during growth. Although RNA levels have been extensively studied in relation to the GR in many organisms, synthesis and degradation rates have, however, been much less investigated. In a previous work, we found a positive correlation between RNA polymerase (RNA pol) II transcription and mRNA degradation with GRs in yeast in batch cultures. Here we extend our study under constant growth conditions in a chemostat and find that overall chromatin-associated RNA pol II levels increase in parallel with the GR. This increase appears to involve the accumulation of partially dephosphorylated RNA pol II with a greater tendency to backtracking, which suggests that the GR modifies the phosphorylation state of RNA pol II at the elongation level. RNA pol I also increases its association with chromatin with the GR, which confirms the general dependence of at least RNA pol I and II transcription on the GR.

在自然选择过程中，细胞必须适应不断变化的环境条件，以保持其适应性，并与其他基因型竞争。生长速率（GR）是这种竞争的决定性因素，它影响着基因的表达。一些基因的mRNA水平随着gr的增加而增加，而另一些基因的mRNA水平则随着gr的增加而降低。mRNA水平取决于RNA聚合酶II合成和降解率之间的动态平衡。RNA聚合酶I和RNA聚合酶III也受到GR的影响，因为它们转录蛋白质合成机制，以在生长过程中制造增加细胞质量的蛋白质。尽管在许多生物体中，RNA水平与GR的关系已被广泛研究，但对其合成和降解率的研究却少得多。在之前的工作中，我们发现在批量培养的酵母中，RNA聚合酶（RNA pol） II转录和mRNA降解与GRs呈正相关。在此，我们在趋化器中持续生长的条件下扩展了我们的研究，发现染色质相关的RNA pol II水平与GR平行增加。这种增加似乎涉及部分去磷酸化的RNA pol II的积累，并且更倾向于回溯，这表明GR在延伸水平上改变了RNA pol II的磷酸化状态。RNA pol I也增加了其与GR中染色质的关联，这证实了至少RNA pol I和II的转录对GR的普遍依赖。

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引用次数: 0

Insights into the target-directed miRNA degradation mechanism in Drosophila ovarian cell culture 果蝇卵巢细胞培养中靶向miRNA降解机制的研究

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-05-04 DOI: 10.1016/j.bbagrm.2025.195092

Natalia Akulenko , Elena Mikhaleva , Sofya Marfina , Ivan Kutelev , Dmitry Kornyakov , Vlad Bobrov , Andrei Artamonov , Georgij Arapidi , Victoria Shender , Sergei Ryazansky

Target-directed miRNA degradation (TDMD) is a process of post-transcriptional regulation of miRNA stability in animals induced by an extended pairing of Ago-bound miRNAs with specialized complementary RNA targets. As suggested by studies on human cell culture, Ago engaged with the extended duplex is recognized by the ZSWIM8 receptor of the Cullin-RING-ligase complex (CRL3), which also contains Cul3, EloB, and EloC proteins. The CRL activity is accelerated by the neddylation of Cul3 with the involvement of the E2 conjugating protein UbcE2M. The CRL ubiquitinates Ago, resulting in proteolysis of Ago and degradation of the released miRNAs. To date, the molecular mechanism of TDMD has not been studied in other species. To further characterize TDMD in animals, we investigated the protein Dora, the Drosophila ortholog of ZSWIM8, in the culture of Drosophila ovarian somatic cells (OSC). We showed that Dora in OSCs localizes in protein granules unrelated to P- and GW-bodies. The dora knockout resulted in the accumulation of multiple miRNAs, including miR-7-5p, and transcriptome-wide affected the mRNA targets of differentially expressed miRNAs. We also showed that Dora associates with proteins of the CRL3 complex, and the depletion of CRL3 components or inhibition of Cul3 neddylation upregulates miR-7-5p. We concluded that the molecular mechanism of TDMD is conserved in humans and Drosophila. Finally, we found that cells without Dora have an impaired Notch signaling pathway, indicating that TDMD in OSCs may contribute to the modulation of the Notch pathway.

靶定向miRNA降解（Target-directed miRNA degradation， TDMD）是通过ago结合miRNA与特异性互补RNA靶标的扩展配对，诱导动物miRNA稳定性的转录后调控过程。人类细胞培养的研究表明，与扩展双工结合的Ago被Cullin-RING-ligase复合物（CRL3）的ZSWIM8受体识别，该复合物还含有Cul3、EloB和EloC蛋白。在E2偶联蛋白UbcE2M的参与下，Cul3的类化修饰加速了CRL的活性。CRL泛素化Ago，导致Ago的蛋白水解和释放的mirna的降解。迄今为止，尚未在其他物种中研究TDMD的分子机制。为了进一步表征动物中的TDMD，我们在果蝇卵巢体细胞（OSC）培养中研究了ZSWIM8的果蝇同源蛋白Dora。我们发现Dora在OSCs中定位于与P-和gw -体无关的蛋白质颗粒中。dora基因敲除导致包括miR-7-5p在内的多个mirna的积累，并且转录组范围内影响差异表达mirna的mRNA靶标。我们还发现Dora与CRL3复合体的蛋白相关，CRL3成分的缺失或Cul3类化修饰的抑制可上调miR-7-5p。我们认为TDMD的分子机制在人类和果蝇中是保守的。最后，我们发现没有Dora的细胞Notch信号通路受损，表明OSCs中的TDMD可能参与了Notch通路的调节。

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引用次数: 0

Navigating the tumor landscape: VEGF, MicroRNAs, and the future of cancer treatment 导航肿瘤景观：VEGF， microrna和癌症治疗的未来

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-05-03 DOI: 10.1016/j.bbagrm.2025.195091

K.P. Ameya, P.P. Ashikha Shirin Usman, Durairaj Sekar

Cancer progression is a multifaceted process influenced by complex interactions within the tumor microenvironment (TME). Central to these dynamics are Vascular Endothelial Growth Factor (VEGF) signalling and microRNA (miRNA) modulation, both of which play critical roles in tumor growth and angiogenesis. VEGF is essential for promoting blood vessel formation; however, its splice variant, VEGF165b, acts as an anti-angiogenic factor, presenting a paradox challenging conventional cancer therapies. Meanwhile, miRNAs regulate gene expression that significantly impacts tumor behaviour by targeting various mRNAs involved in signalling pathways. The interplay between VEGF and miRNAs opens new avenues for targeted therapies designed to disrupt the networks supporting tumor growth. Additionally, the concept of exploiting the unique properties of VEGF splice variants is being explored to develop novel treatments that enhance anti-angiogenic effects while minimizing side effects. Understanding this is crucial for advancing personalized therapies that can effectively address the challenges posed by tumor adaptability and resistance mechanisms.

癌症的进展是一个多方面的过程，受肿瘤微环境（TME）内复杂相互作用的影响。这些动态的核心是血管内皮生长因子（VEGF）信号传导和microRNA （miRNA）调节，两者在肿瘤生长和血管生成中起着关键作用。VEGF对促进血管形成至关重要；然而，它的剪接变体VEGF165b作为一种抗血管生成因子，提出了一个挑战传统癌症治疗的悖论。同时，miRNAs通过靶向参与信号通路的各种mrna来调节基因表达，显著影响肿瘤行为。VEGF和mirna之间的相互作用为靶向治疗开辟了新的途径，旨在破坏支持肿瘤生长的网络。此外，利用VEGF剪接变体的独特特性的概念正在被探索，以开发新的治疗方法，增强抗血管生成作用，同时最大限度地减少副作用。了解这一点对于推进个性化治疗至关重要，可以有效地解决肿瘤适应性和耐药机制带来的挑战。

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引用次数: 0

The crucial role of small heat shock proteins in prostate cancer: mechanisms and new therapeutic perspectives 小热休克蛋白在前列腺癌中的关键作用：机制和新的治疗前景

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-04-11 DOI: 10.1016/j.bbagrm.2025.195090

Yuankang Feng , Jialu Ma , Zhihao Bo , Dan Yue , Yong Wang

As resistance to new anti-androgen drugs occurs more frequently, increasing numbers of researchers are exploring alternative key molecular targets for prostate cancer treatment. The small heat shock protein (sHSP) family is a subclass of heat shock proteins (HSPs). Due to the smaller molecular size of their monomers, they often function as large oligomeric complexes with diverse biological roles, thus garnering increasing attention from urologists. Different members of the sHSP family exhibit distinct biological roles in prostate cancer, offering a new perspective for precision therapy. In this review, we summarize the specific roles of sHSP family members in prostate cancer and analyze their similarities and differences. Additionally, we discuss and review the drugs targeting various sHSPs in prostate cancer, providing new insights into the exploration and further application of sHSP-targeted therapies.

随着新型抗雄激素药物耐药性的频繁发生，越来越多的研究人员正在探索前列腺癌治疗的其他关键分子靶点。小热休克蛋白（sHSP）家族是热休克蛋白的一个亚类。由于其单体的分子尺寸较小，它们通常作为具有多种生物学作用的大型寡聚复合物发挥作用，因此越来越受到泌尿科医生的关注。sHSP家族的不同成员在前列腺癌中表现出不同的生物学作用，为精准治疗提供了新的视角。本文就sHSP家族成员在前列腺癌中的具体作用进行综述，并分析其异同点。此外，我们还讨论和回顾了针对各种shsp在前列腺癌中的药物，为shsp靶向治疗的探索和进一步应用提供了新的见解。

引用次数: 0

Genome-wide screening reveals repression by nuclear exosome as a prerequisite for intron-mediated enhancement in Saccharomyces cerevisiae 全基因组筛选显示核外泌体的抑制是酿酒酵母内含子介导的增强的先决条件

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-04-10 DOI: 10.1016/j.bbagrm.2025.195089

Hiroki Kikuta , Shunya Takeda , Rinji Akada , Hisashi Hoshida

Introns can enhance gene expression, a phenomenon called intron-mediated enhancement (IME). Previously proposed IME mechanisms do not sufficiently explain the variability in enhancement levels, suggesting that IME mechanism has not been fully understood. A comprehensive screening of genes involved in IME can provide valuable insights. Recently, using a luciferase coding sequence (yCLuc), we showed that IME functions by relieving repression rather than simply enhancing expression. The expression of yCLuc is repressed by the specific nucleotide sequence UCUU, and adding an intron relieves this repression in the yeast Saccharomyces cerevisiae. Herein, genome-wide screenings were conducted using S. cerevisiae knockout strain libraries to identify genes involved in IME. For screening, yCLuc was expressed with and without an intron in knockout strains. Consequently, CDC73, a regulator of RNA polymerase II (RNAPII), was identified as essential for enhancement. Additionally, 23 genes specifically involved in the repression were identified. These 23 genes are related to nuclear exosomes, RNA modification, RNAPII regulation, the nuclear pore complex, ribosomes, and chromatin modification. Among these, genes associated with nuclear exosomes, which degrade various RNAs in the nucleus, showed the largest impact on expression. The RNA sequence UCUU has been reported as a target for RNA degradation by nuclear exosomes. These findings suggested that UCUU-containing coding sequences are primarily repressed via RNA degradation by the nuclear exosome through UCUU recognition, with this repression being relieved by the presence of an intron.

内含子可以增强基因表达，这种现象被称为内含子介导增强（IME）。先前提出的IME机制并不能充分解释增强水平的可变性，这表明IME机制尚未被完全理解。全面筛选与IME有关的基因可以提供有价值的见解。最近，利用荧光素酶编码序列（yCLuc），我们发现IME的功能是缓解抑制，而不仅仅是增强表达。yCLuc的表达受特定核苷酸序列UCUU的抑制，在酿酒酵母中加入内含子可以缓解这种抑制。本文利用酿酒葡萄球菌敲除菌株文库进行全基因组筛选，以鉴定与IME相关的基因。为了筛选，yCLuc在敲除菌株中有和没有内含子表达。因此，RNA聚合酶II （RNAPII）的调节因子CDC73被确定为增强的必要条件。此外，还鉴定了23个特异性参与抑制的基因。这23个基因与核外泌体、RNA修饰、RNAPII调控、核孔复合体、核糖体和染色质修饰有关。其中，与核外泌体相关的基因对表达的影响最大，核外泌体降解细胞核中的各种rna。RNA序列UCUU已被报道为核外泌体RNA降解的靶标。这些发现表明，含有UCUU的编码序列主要通过RNA降解被核外泌体通过UCUU识别抑制，而这种抑制被内含子的存在所缓解。

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引用次数: 0

Specific DNA features of the RNA polymerase I core promoter element targeted by core factor 核心因子所针对的 RNA 聚合酶 I 核心启动子元件的特定 DNA 特征

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-04-09 DOI: 10.1016/j.bbagrm.2025.195088

Nathan J. Munoff, Brian J. Zeberl, Matthew A. Palmer, Wayne A. Decatur, Bridget M. Walker, Jyoti D. Adala, Zsuzsa K. Szemere, Aula M. Fakhouri, Bruce A. Knutson

RNA polymerase I (Pol I) is essential for ribosomal RNA (rRNA) synthesis, driving ribosome biogenesis in eukaryotes. Transcription initiation by Pol I requires core factor (CF) binding to the core element (CE) of the ribosomal DNA (rDNA) promoter. Despite structural conservation across species, significant sequence variability suggests CF recognizes DNA through structural features rather than specific sequences. We investigated CF's DNA binding preferences to elucidate the role of DNA structural properties in CE recognition. Analysis of CE sequences from 35 fungal species revealed conserved structural features, notably a rigid AT-rich patch at positions −22 to −20 and a conserved G base pair at position −24. Competition-based electrophoretic mobility shift assays (EMSA) with single base-pair substitutions showed CF tolerates mutations at many positions but is sensitive to changes in the AT-rich patch. Loss of CF binding correlated with alterations in DNA structural properties such as increased bendability, decreased curvature, widened minor groove width, and altered helix twist. In vitro SELEX experiments identified novel CE sequences preferentially bound by CF, exhibiting increased GC content, higher bendability, and decreased curvature despite lacking sequence conservation. Classification based on bendability profiles revealed CF preferentially binds bendable sequences. In vivo selection assays confirmed these findings, demonstrating consistent CF binding preferences within a cellular context. Our results indicate that CF recognizes and binds to the CE primarily through specific DNA structural features rather than nucleotide sequences. Structural properties like bendability, curvature, and minor groove width are critical determinants of CF binding, facilitating effective Pol I transcription initiation.

RNA聚合酶I （Pol I）是真核生物中核糖体RNA （rRNA）合成和驱动核糖体生物发生所必需的。Pol I的转录起始需要核心因子（CF）与核糖体DNA （rDNA）启动子的核心元件（CE）结合。尽管物种之间存在结构守恒，但显著的序列差异表明CF通过结构特征而不是特定序列识别DNA。我们研究了CF的DNA结合偏好，以阐明DNA结构特性在CE识别中的作用。对35种真菌CE序列的分析显示，CE序列具有保守的结构特征，特别是在- 22 ~ - 20位置有一个刚性的at -rich patch，在- 24位置有一个保守的G碱基对。采用单碱基对替换的基于竞争的电泳迁移迁移试验（EMSA）显示，CF可以耐受许多位置的突变，但对富含at的斑块的变化敏感。CF结合的丧失与DNA结构特性的改变相关，如可弯曲性增加、曲率降低、小凹槽宽度变宽和螺旋扭曲改变。体外SELEX实验发现，新的CE序列优先与CF结合，尽管缺乏序列保守性，但GC含量增加，可弯曲性提高，曲率降低。基于可弯曲性特征的分类显示，CF优先结合可弯曲序列。体内选择分析证实了这些发现，证明了在细胞背景下一致的CF结合偏好。我们的研究结果表明，CF主要通过特定的DNA结构特征而不是核苷酸序列来识别和结合CE。结构特性，如可弯曲性，曲率和小凹槽宽度是CF结合的关键决定因素，促进有效的Pol I转录起始。

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引用次数: 0

Hypoxia inducible factor HIF1α elevates expression of mRNA capping enzyme during cobalt chloride-induced hypoxia 缺氧诱导因子HIF1α在氯化钴诱导的缺氧过程中升高mRNA capping酶的表达。

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-04-05 DOI: 10.1016/j.bbagrm.2025.195087

Safirul Islam, Chandrama Mukherjee

In response to hypoxia, hypoxia-inducible factors (HIFs) control the transcriptomic output to mitigate the hypoxic stress. Long noncoding RNAs (lncRNA) are found to be very crucial in regulating hypoxia. Like mRNAs, lncRNAs are protected by 5′ caps that are added by mRNA capping enzyme (CE) in the nucleus. The previous concept that capping takes place in the nucleus was changed by the recognition of a cytoplasmic pool of capping enzyme (cCE). cCE has been shown to recap its substrate uncapped mRNAs or long noncoding RNAs (lncRNAs) present in the cytoplasm, preventing their degradation, even during arsenite-induced oxidative stress. In this study, we examined the effect of CoCl₂ induced hypoxia on cCE and its function in regulating the substrate lncRNAs.

Here, we show that CoCl₂ induced hypoxia elevates the expressions of nuclear and cytoplasmic CE in HIF1α dependent manner as evidenced by Chromatin immunoprecipitation and HIF1α inhibitor experiments. Furthermore, we found cCE post-transcriptionally controls the stability of its target lncRNAs amidst CoCl₂ induced hypoxia. These results suggest that cCE, upregulated by HIF1α, may act as a posttranscriptional modulator for a few cCE-targeted lncRNAs.

低氧诱导因子（hif）通过调控转录组输出来缓解低氧胁迫。长链非编码rna （Long noncoding rna, lncRNA）在缺氧调控中起着至关重要的作用。与mRNA一样，lncRNAs受到细胞核中mRNA capping酶（CE）添加的5'帽的保护。由于认识到胞质盖顶酶（cCE）的存在，以前认为盖顶发生在细胞核内的观念发生了改变。研究表明，cCE可以重新封装存在于细胞质中的底物无帽mrna或长链非编码rna (lncRNAs)，即使在亚砷酸盐诱导的氧化应激过程中也能阻止它们的降解。在本研究中，我们研究了CoCl2诱导的缺氧对cCE的影响及其在调节底物lncrna中的功能。在这里，我们通过染色质免疫沉淀和HIF1α抑制剂实验证明，CoCl2诱导的缺氧以依赖于HIF1α的方式提高细胞核和细胞质CE的表达。此外，我们发现cCE通过转录后调控其靶lncrna在CoCl2诱导的缺氧中的稳定性。这些结果表明，hif - 1α上调的cCE可能作为一些靶向cCE的lncrna的转录后调节剂。

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引用次数: 0

Transcription factors associated with regulation of transcriptome in human thigh and calf muscles at baseline and after six days of disuse 在基线和停用6天后，与人大腿和小腿肌肉转录组调节相关的转录因子。

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-02-19 DOI: 10.1016/j.bbagrm.2025.195086

Anna A. Borzykh , Pavel A. Makhnovskii , Ivan I. Ponomarev, Tatiana F. Vepkhvadze, Egor M. Lednev, Ilya V. Rukavishnikov, Oleg I. Orlov, Elena S. Tomilovskaya, Daniil V. Popov

Disuse has a negative impact on the postural muscles of the trunk and legs. Different leg muscles demonstrate a differentiated and conservative response to disuse, in terms of a decrease in muscle mass, strength, aerobic performance, and changes in gene expression. We aimed to identify transcription factors regulating gene expression at baseline and after disuse in human m. soleus – a “slow” muscle with a strong postural function, and “mixed” m. vastus lateralis. Biopsies were taken from these muscles prior to and after 6 days of strict disuse (dry immersion). The enriched transcription factor binding sites (and corresponding factors) in the individual promoter regions of co-expressed genes were examined using the positional weight matrix approach. The baseline transcriptomic profiles and the disuse-induced changes (RNA-seq) differ significantly between muscles. In particular, the specific and significant response to disuse in m. soleus was found to be strongly related to the suppression of genes regulating the mitochondrial energy metabolism, the activation of the inflammatory response and the ubiquitin-proteasome system. This response is associated with the proinflammatory transcription factors such as families IRF, STAT, and other. The validity of approximately two-thirds of the predicted transcription factors was indirectly confirmed by the analysis of their function described in the literature. These identified transcription factors appear to be promising candidates for future targeted studies that mechanistically investigate gene expression regulation in various muscles at baseline, following disuse or inactivity.

不使用对躯干和腿部的姿势肌肉有负面影响。在肌肉质量、力量、有氧运动表现和基因表达变化方面，不同的腿部肌肉表现出不同的和保守的废用反应。我们的目的是确定在人类比目鱼肌（一种具有强大姿势功能的“慢”肌肉）和“混合”股外侧肌中基线和废用后调节基因表达的转录因子。在严格弃用（干浸泡）之前和之后6 天，对这些肌肉进行活组织检查。利用位置权重矩阵法检测共表达基因的单个启动子区域中富集的转录因子结合位点（和相应的因子）。基线转录组谱和废用诱导的变化（RNA-seq）在肌肉之间有显著差异。特别是，比目鱼对废用的特异性和显著反应被发现与调节线粒体能量代谢的基因的抑制、炎症反应的激活和泛素-蛋白酶体系统的激活密切相关。这种反应与促炎转录因子如家族IRF、STAT等有关。大约三分之二的预测转录因子的有效性通过文献中描述的功能分析间接证实。这些已确定的转录因子似乎是未来有针对性的研究的有希望的候选者，这些研究将在基线、不使用或不活动后对各种肌肉的基因表达调控进行机制研究。

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引用次数: 0

UPS regulation of gene expression and genome integrity UPS对基因表达和基因组完整性的调控。

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-01-30 DOI: 10.1016/j.bbagrm.2025.195078

Sukesh R. Bhaumik

引用次数: 0

TAP-MS analysis of FACT interactions and regulation by a ubiquitin ligase, San1 泛素连接酶San1对FACT相互作用及调控的TAP-MS分析。

IF 2.6 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et Biophysica Acta-Gene Regulatory Mechanisms

Pub Date : 2025-01-22 DOI: 10.1016/j.bbagrm.2025.195077

Priyanka Barman, Pritam Chakraborty, Shalini Guha, Amala Kaja, Rhea Bhaumik, Sukesh R. Bhaumik

An evolutionarily conserved heterodimeric FACT (Facilitates chromatin transcription) regulates transcription, DNA repair, replication and other cellular processes via its interactions with other proteins. FACT is recently found to be regulated via ubiquitylation and 26S proteasomal degradation, alteration of which is associated with aberrant transcription and genome integrity. However, there has not been a systematic study to analyze FACT interactions proteome-wide in the presence and absence of its UPS (Ubiquitin-proteasome system) regulation, which could reveal new FACT interactors with mechanistic and functional implications. Here, we have adopted a proteome-wide approach via TAP (Tandem affinity purification)-mediated pull-down of FACT and its interactors from the soluble and insoluble cellular fractions followed by MS (Mass-spectrometry) analysis. We find distinct interactors of FACT in the soluble and insoluble fractions in addition to a common set in both. While a set of all these interactors overlaps with previously known FACT partners, many are new, which are involved in different cellular processes such as transcription, DNA repair and chromatin regulation. Further, an intrinsically disordered ubiquitin ligase, San1, that ubiquitylates the Spt16 component of FACT for proteasomal degradation to regulate chromatin, transcription and genome integrity is found to influence the interactions of FACT with a set of proteins including epigenetic, transcription and DNA repair factors. Collectively, our results unveil proteome-wide FACT interactions and regulation by a ubiquitin ligase, hence shedding much light on FACT networks with functional and mechanistic implications.

一种进化保守的异源二聚体 FACT（促进染色质转录）通过与其他蛋白质的相互作用调节转录、DNA 修复、复制和其他细胞过程。最近发现 FACT 可通过泛素化和 26S 蛋白质体降解进行调控，而泛素化和 26S 蛋白质体降解的改变与转录异常和基因组完整性有关。然而，目前还没有系统的研究来分析在存在和不存在 UPS（泛素-蛋白酶体系统）调控的情况下 FACT 与整个蛋白体的相互作用，这可能会揭示出具有机制和功能意义的新的 FACT 相互作用因子。在这里，我们采用了一种全蛋白质组的方法，通过串联亲和纯化（TAP）介导，从可溶性和非可溶性细胞组分中提取 FACT 及其相互作用物，然后进行质谱分析。我们在可溶性和不可溶性馏分中发现了不同的 FACT 相互作用体，此外还在两者中发现了一组共同的相互作用体。所有这些相互作用者中有一部分与以前已知的 FACT 伙伴重叠，但也有许多是新的，它们参与了转录、DNA 修复和染色质调控等不同的细胞过程。此外，我们还发现了一种内在无序泛素连接酶 San1，它能泛素化 FACT 的 Spt16 成分，使其蛋白酶体降解，从而调节染色质、转录和基因组的完整性。总之，我们的研究结果揭示了整个蛋白质组的 FACT 相互作用以及泛素连接酶的调控，从而揭示了 FACT 网络的功能和机理。

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引用次数: 0