Germline mutations in the folliculin (FLCN) tumor suppressor gene are linked to Birt-Hogg-Dubé (BHD) syndrome, a dominantly inherited genetic disease characterized by predisposition to fibrofolliculomas, lung cysts, and renal cancer. Most BHD-linked FLCN variants include large deletions and splice site aberrations predicted to cause loss of function. The mechanisms by which missense variants and short in-frame deletions in FLCN trigger disease are unknown. Here, we present an integrated computational and experimental study that reveals that the majority of such disease-causing FLCN variants cause loss of function due to proteasomal degradation of the encoded FLCN protein, rather than directly ablating FLCN function. Accordingly, several different single-site FLCN variants are present at strongly reduced levels in cells. In line with our finding that FLCN variants are protein quality control targets, several are also highly insoluble and fail to associate with the FLCN-binding partners FNIP1 and FNIP2. The lack of FLCN binding leads to rapid proteasomal degradation of FNIP1 and FNIP2. Half of the tested FLCN variants are mislocalized in cells, and one variant (ΔE510) forms perinuclear protein aggregates. A yeast-based stability screen revealed that the deubiquitylating enzyme Ubp15/USP7 and molecular chaperones regulate the turnover of the FLCN variants. Lowering the temperature led to a stabilization of two FLCN missense proteins, and for one (R362C), function was re-established at low temperature. In conclusion, we propose that most BHD-linked FLCN missense variants and small in-frame deletions operate by causing misfolding and degradation of the FLCN protein, and that stabilization and resulting restoration of function may hold therapeutic potential of certain disease-linked variants. Our computational saturation scan encompassing both missense variants and single site deletions in FLCN may allow classification of rare FLCN variants of uncertain clinical significance.

滤泡素 ( FLCN ) 肿瘤抑制基因的种系突变与 Birt-Hogg-Dubé (BHD) 综合征有关，这是一种显性遗传性疾病，其特征是易患纤维毛囊瘤、肺囊肿和肾癌。大多数 BHD 连接的FLCN变体包含大量缺失和剪接位点畸变，预计会导致功能丧失。 FLCN中错义变异和短框内缺失引发疾病的机制尚不清楚。在这里，我们提出了一项综合的计算和实验研究，揭示了大多数此类引起疾病的FLCN变异由于编码的 FLCN 蛋白的蛋白酶体降解而导致功能丧失，而不是直接消除 FLCN 功能。因此，细胞中存在的几种不同的单位点 FLCN 变体的水平大大降低。根据我们的发现，FLCN 变体是蛋白质质量控​​制目标，其中一些变体也是高度不溶的，并且无法与 FLCN 结合伙伴 FNIP1 和 FNIP2 结合。 FLCN 结合的缺乏导致 FNIP1 和 FNIP2 的快速蛋白酶体降解。一半的测试 FLCN 变体在细胞中错误定位，其中一种变体 (ΔE510) 形成核周蛋白聚集体。基于酵母的稳定性筛选显示去泛素化酶 Ubp15/USP7 和分子伴侣调节 FLCN 变体的周转。降低温度导致两种 FLCN 错义蛋白稳定，其中一种 (R362C) 在低温下重新建立了功能。 总之，我们提出，大多数 BHD 连锁的FLCN错义变体和小的框内缺失是通过引起 FLCN 蛋白的错误折叠和降解来起作用的，并且稳定和由此产生的功能恢复可能具有某些与疾病相关的变体的治疗潜力。我们的计算饱和扫描涵盖FLCN中的错义变异和单位点缺失，可能允许对临床意义不确定的罕见 FLCN 变异进行分类。