A key but often overlooked step in gene regulation is the processing of precursor messenger RNAs, the molecules that first emerge from the transcription of a gene. These pre-mRNAs contain sequences (exons) encoding information required to make a protein, interspersed with non-coding regions (introns) that must typically be removed before protein production can start. A large molecular machine, the spliceosome, distinguishes introns from exons, removes the former and joins the latter to create a mature mRNA template.
基因调控中一个关键但常被忽视的环节是前体信使RNA的加工过程——这是基因转录后最先生成的分子。这些前体mRNA包含编码蛋白质信息的外显子序列，其间穿插着需要在蛋白质合成前去除的非编码区域（内含子）。剪接体这一大型分子机器能够识别内外显子边界，去除内含子并连接外显子形成成熟的mRNA模板。
Cells can fine-tune these complex splicing events to control what proteins are made, when, and in what form (Boutz et al., 2015). Thus, splicing allows the organism to meet changing demands quickly and flexibly. Deliberately leaving in "detained introns" prevents target pre-mRNAs from being exported from the nucleus, which allows the cell to delay or prevent the production of certain proteins without degrading the associated RNA transcripts (Yap et al., 2012). Exon skipping, on the other hand, occurs when the spliceosome skips an exon to allow an mRNA to be produced albeit with an altered coding sequence. "Decoy" exons also occur. Although their mode of action remains unclear, these exons contained within introns are believed to recruit and then "stall" the spliceosome, preventing it from proceeding with the normal splicing process.
细胞可通过精细调控这些复杂的剪接事件来决定蛋白质的合成时序与形式（Boutz等，2015）。这种机制使生物能快速灵活地应对变化需求。刻意保留的"滞留内含子"可阻止靶向pre-mRNA输出细胞核，使细胞能在不降解RNA转录本的情况下延迟或抑制特定蛋白质生成（Yap等，2012）。当剪接体跳过外显子时会发生外显子跳跃，虽产生mRNA但其编码序列已改变。还存在"诱饵外显子"现象——尽管作用机制尚不明确，这类内含子中的外显子被认为能招募并"阻滞"剪接体，干扰正常剪接进程。
Recent work has shown that a highly dynamic protein modification, known as O-GlcNAc, plays a key role in regulating detained intron splicing (Tan et al., 2020). O-GlcNAcylation consists of the addition of a small sugar molecule (GlcNAc) onto certain amino acids, which can alter the activity and location of thousands of proteins in a cell. It helps modulate gene expression and many crucial signaling pathways, such as those involved in responding to DNA damage or maintaining cell identity in early development (Bond and Hanover, 2015; Zachara et al., 2022; Fehl and Hanover, 2022). O-GlcNAc levels vary in response to broader environmental signals, in particular stressors or variations in nutrient availability. As such, this process allows cells to adjust their response and retain their internal balance in the face of ever-changing conditions. Finely regulating O-GlcNAcylation is therefore crucial for survival, with deregulation being linked to some forms of X-linked intellectual disability (Konzman et al., 2020; Vaidyanathan et al., 2017).
最新研究显示，一种高度动态的蛋白质翻译后修饰——O-GlcNAc修饰在调控内含子滞留中起关键作用（Tan等，2020）。O-GlcNAc糖基化修饰是指将N-乙酰葡萄糖胺（GlcNAc）添加至特定氨基酸的修饰过程，这种修饰能改变细胞中数千种蛋白质的活性与定位。它参与调控基因表达和多个关键信号通路，如DNA损伤应答、早期发育中细胞身份维持等（Bond与Hanover，2015；Zachara等，2022；Fehl与Hanover，2022）。O-GlcNAc水平会随环境信号（特别是应激源或营养物质变化）动态调整，使细胞在不断变化的环境中维持内稳态。精准调控O-GlcNAc修饰对生存至关重要，其失调与某些X染色体连锁智力障碍疾病相关（Konzman等，2020；Vaidyanathan等，2017）。
Interestingly, the control of O-GlcNAcylation itself seems to be linked to detained intron splicing (Tan et al., 2020). Two enzymes, O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), respectively add and remove GlcNAc to/from proteins (Bond and Hanover, 2015; Zachara et al., 2022). An intricate feedback loop maintains stability in the system: when O-GlcNAc levels rise, for instance, OGT production decreases while that of OGA increases. More precisely, under high O-GlcNAc levels, introns are retained in the pre-mRNA transcript of the OGT gene, preventing protein expression. A similar mechanism unfolds for OGA when O-GlcNAc is low (Figure 1A and B; Park et al., 2017; Tan et al., 2020). Should this fail to restore healthy levels of O-GlcNAc, the cell responds by specifically changing detained intron splicing across the whole genome. What controls this O-GlcNAc-driven splicing regulation, however, is still poorly understood. Now, in eLife, Ashwin Govindan and Nicholas Conrad from the University of Texas Southwestern Medical Center report having identified the splicing factor SFSWAP as a key player required for this process (Figure 1C; Govindan and Conrad, 2025).
有趣的是，O-GlcNAc修饰本身的调控也与内含子滞留存在关联（Tan等，2020）。O-GlcNAc转移酶（OGT）和O-GlcNAc酶（OGA）分别负责将GlcNAc添加到蛋白上或将其去除（Bond与Hanover，2015；Zachara等，2022）。该系统通过精巧的反馈环维持稳定：例如当O-GlcNAc水平升高时，OGT生成减少而OGA增加。更精确地说，高O-GlcNAc水平下，OGT基因pre-mRNA会保留内含子从而抑制蛋白表达；当O-GlcNAc水平低时，OGA基因呈现类似机制（图1A和B；Park等，2017；Tan等，2020）。当这种机制无法恢复O-GlcNAc的健康水平时，细胞会通过改变全基因组范围的内含子滞留作出响应。但这种O-GlcNAc驱动的剪接调控机制尚不清楚。德州大学西南医学中心的Ashwin Govindan和Nicholas Conrad在eLife发表的研究中鉴定出剪接因子SFSWAP是该过程的关键调控者（图1C；Govindan与Conrad，2025）。
Figure 1
SFSWAP as a regulator of O-GlcNAc through splicing.
O-GlcNAc is a protein modification that acts as an important regulator of epigenetics and signaling. The amount of O-GlcNAcylation within a cell changes in response to the environment, in particular...
The team focused on the gene coding for OGT, devising an elegant screening strategy to identify the molecular actors that contribute to its splicing under O-GlcNAc control. They first created an OGT-based genetic construct that appropriately responded to O-GlcNAc levels, and whose detained intron splicing could easily be detected. This reporter was formed of the sequence coding for the fluorescent protein GFP, in which intron 4 from OGT was introduced (alongside the corresponding exons 4 and 5); if this intron was retained, GFP production dropped. Intron 4 was chosen in part because it contains a "decoy exon", recruiting spliceosomes but blocking normal splicing (Parra et al., 2018).
研究团队聚焦于OGT基因，设计了一种创新的筛选策略来识别参与O-GlcNAc调控下剪接过程的分子因素。他们构建了对O-GlcNAc水平敏感的OGT基因报告系统，该系统可通过滞留内含子剪接状态的变化被检测。该报告系统由表达荧光蛋白GFP的序列构成，在其中插入了OGT的第4内含子（包含相邻的第4和第5外显子）；当该内含子滞留时，GFP表达量下降。选择第4内含子部分原因在于其包含"诱饵外显子"，能招募剪接体但阻断正常剪接（Parra等，2018）。
Govindan and Conrad introduced their construct in human cell lines in which they systematically deleted genes one at a time. The cells were then manipulated so that their O-GlcNAc levels increased or decreased, and GFP expression was carefully monitored. A range of genes emerged as potentially controlling OGT splicing under these conditions, including several that code for components of the spliceosome. One particularly promising candidate was then selected based on rigorous criteria: SFSWAP, a splicing factor whose equivalent in Drosophila regulates alternative splicing in various transcripts, including its own. In humans, SFSWAP is known to control the splicing of several genes (including Tau, CD45 and fibronectin) by inhibiting the inclusion of specific exons.
Govindan和Conrad在人类细胞系中引入该构建体，并系统性地进行逐个基因敲除。通过调节细胞O-GlcNAc水平变化并持续监测GFP表达，筛选出一系列可能调控OGT剪接的候选基因，包括多个编码剪接体组分的基因。根据严格筛选标准，最终确定SFSWAP为关键候选因子——这是一种剪接因子，在果蝇中的同源物调控包括自身在内的多个转录本的可变剪接。在人类中，SFSWAP已知通过抑制特定外显子包含来调控多个基因（包括Tau、CD45和纤维连接蛋白）的剪接。
Next, Govindan and Conrad investigated how SFSWAP contributed to five types of splicing events (detained introns, skipped exons, alternate 5' splice sites, alternate 3' splice sites and mutually exclusive exons) under various O-GlcNAc levels. Overall, the analyses show that SFSWAP acts on two of these mechanisms – detained introns and skipped exons – and that it promotes the retention of introns in a wide range of transcripts.
随后，Govindan和Conrad研究了SFSWAP在不同O-GlcNAc水平下对五类剪接事件（滞留内含子、外显子跳跃、可变5'剪接位点、可变3'剪接位点和互斥外显子）的影响。分析表明，SFSWAP主要作用于滞留内含子和外显子跳跃两类机制，且其促进内含子滞留的效应存在于多种转录本中。
Additional experiments examining the impact of SFSWAP on pre-mRNAs with or without decoy exons led Govindan and Conrad to propose that this factor increases intron retention by acting at a later stage in the splicing cycle. According to this model, SFSWAP may interfere with how the spliceosome proceeds to excise certain sequences, but not how this machinery assembles at specific locations on the transcript. Taken together, these findings have important implications for understanding the complex interplay between environmentally responsive O-GlcNAc metabolism and the global regulation of splicing – uncovering a feedback loop by which global changes can directly influence gene expression at a post-transcriptional level.
通过比较SFSWAP对含/不含诱饵外显子pre-mRNA的影响，他们提出SFSWAP通过作用于剪接循环后期阶段来增强内含子滞留的机制模型。根据该模型，SFSWAP可能干扰剪接体对特定序列的切除进程，但不影响其在转录本特定位置的组装。这些发现揭示了环境响应性O-GlcNAc代谢与全局剪接调控之间复杂的相互作用，发现了一个通过转录后水平直接调控基因表达的反馈环路。
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