Germline nonsense and canonical splice site variants identified in disease-causing genes are generally considered as loss-of-function (LoF) alleles and classified as pathogenic. However, a fraction of such variants could maintain function through their impact on RNA splicing. To test this hypothesis, we used the alternatively spliced BRCA2 exon 12 (E12) as a model system since its in-frame skipping leads to a potentially functional protein. All E12 variants corresponding to putative LoF variants or predicted to alter splicing (n=40) were selected from human variation databases and characterized for their impact on splicing in minigene assays and, when available, in patient lymphoblastoid cell lines. Moreover, a selection of variants was analyzed in a mouse embryonic stem cell-based functional assay. Using these complementary approaches, we demonstrate that a subset of variants, including nonsense variants, induced in-frame E12 skipping through the modification of splice sites or regulatory elements and, consequently, led to an internally deleted but partially functional protein. These data provide evidence for the first time in a cancer-predisposition gene, that certain presumed null variants can retain function due to their impact on splicing. Further studies are required to estimate cancer-risk associated with these hypomorphic variants. More generally, our findings highlight the need to exercise caution in the interpretation of putative LoF variants susceptible to induce in-frame splicing modifications.

中文翻译：

---

跳过废话以维持功能：BRCA2外显子12的范例。

在致病基因中鉴定的种系废话和规范剪接位点变异通常被认为是功能丧失（LoF）等位基因，并被归类为致病性。但是，这类变体的一部分可以通过其对RNA剪接的影响来维持功能。为了验证这一假设，我们使用了选择性剪接的BRCA2外显子12（E12）作为模型系统，因为其框内跳跃会导致潜在的功能蛋白。从人类变异数据库中选择与推定的LoF变异相对应或预计会改变剪接的所有E12变异（n = 40），并在微基因测定以及患者淋巴母细胞样细胞系中表征其对剪接的影响。而且，在基于小鼠胚胎干细胞的功能测定中分析了变体的选择。使用这些补充方法，我们证明了变体的一个子集，包括无意义的变体，通过剪接位点或调控元件的修饰而诱导了框内E12的跳跃，因此导致了内部缺失但部分功能的蛋白质。这些数据首次在癌症易感基因中提供了证据，即某些假定的无效变体由于其对剪接的影响而可以保留功能。需要进一步的研究来估计与这些亚型变异相关的癌症风险。更普遍地说，我们的发现强调在解释容易诱导框内剪接修饰的推定LoF变异体时，必须谨慎行事。导致内部缺失但部分功能的蛋白质。这些数据首次在癌症易感基因中提供了证据，即某些假定的无效变体由于其对剪接的影响而可以保留功能。需要进一步的研究来估计与这些亚型变异相关的癌症风险。更一般而言，我们的发现突出了在解释易于诱发框内剪接修饰的推定LoF变异体时需要谨慎行事的必要性。导致内部缺失但部分功能的蛋白质。这些数据首次在癌症易感基因中提供了证据，即某些假定的无效变体由于其对剪接的影响而可以保留功能。需要进一步的研究来估计与这些亚型变异相关的癌症风险。更普遍地说，我们的发现突出了在解释容易诱导框内剪接修饰的推定LoF变异体时需要谨慎行事的必要性。