RESUMO
Birt-Hogg-Dubé (BHD) syndrome is an autosomal dominant disorder where patients are predisposed to kidney cancer, lung and kidney cysts and benign skin tumors. BHD is caused by heterozygous mutations affecting folliculin (FLCN), a conserved protein that is considered a tumor suppressor. Previous research has uncovered multiple roles for FLCN in cellular physiology, yet it remains unclear how these translate to BHD lesions. Since BHD manifests hallmark characteristics of ciliopathies, we speculated that FLCN might also have a ciliary role. Our data indicate that FLCN localizes to motile and non-motile cilia, centrosomes and the mitotic spindle. Alteration of FLCN levels can cause changes to the onset of ciliogenesis, without abrogating it. In three-dimensional culture, abnormal expression of FLCN disrupts polarized growth of kidney cells and deregulates canonical Wnt signalling. Our findings further suggest that BHD-causing FLCN mutants may retain partial functionality. Thus, several BHD symptoms may be due to abnormal levels of FLCN rather than its complete loss and accordingly, we show expression of mutant FLCN in a BHD-associated renal carcinoma. We propose that BHD is a novel ciliopathy, its symptoms at least partly due to abnormal ciliogenesis and canonical Wnt signalling.
Assuntos
Síndrome de Birt-Hogg-Dubé/fisiopatologia , Cílios/fisiologia , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Sequência de Bases , Síndrome de Birt-Hogg-Dubé/genética , Linhagem Celular , Polaridade Celular , Proliferação de Células , Centrossomo/fisiologia , Cílios/patologia , Humanos , Rim/fisiologia , Microtúbulos/fisiologia , Dados de Sequência Molecular , Análise de Sequência de DNA , Via de Sinalização WntRESUMO
Birt-Hogg-Dubé (BHD) syndrome patients are uniquely susceptible to all renal tumour subtypes. The underlying mechanism of carcinogenesis is unclear. To study cancer development in BHD, we used human proximal kidney (HK2) cells and found that long-term folliculin (FLCN) knockdown was required to increase their tumorigenic potential, forming larger spheroids in non-adherent conditions. Transcriptomic and proteomic analysis uncovered links between FLCN, cell cycle control and the DNA damage response (DDR) machinery. HK2 cells lacking FLCN had an altered transcriptome profile with cell cycle control gene enrichment. G1/S cell cycle checkpoint signaling was compromised with heightened protein levels of cyclin D1 (CCND1) and hyperphosphorylation of retinoblastoma 1 (RB1). A FLCN interactome screen uncovered FLCN binding to DNA-dependent protein kinase (DNA-PK). This novel interaction was reversed in an irradiation-responsive manner. Knockdown of FLCN in HK2 cells caused a marked elevation of γH2AX and RB1 phosphorylation. Both CCND1 and RB1 phosphorylation remained raised during DNA damage, showing an association with defective cell cycle control with FLCN knockdown. Furthermore, Flcn-knockdown C. elegans were defective in cell cycle arrest by DNA damage. This work implicates that long-term FLCN loss and associated cell cycle defects in BHD patients could contribute to their increased risk of cancer.
RESUMO
Birt-Hogg-Dubé syndrome (BHD) is an autosomal dominant condition characterised by the presence of facial fibrofolliculomas, pulmonary cysts which may be associated with spontaneous pneumothorax and renal tumours. Germline mutations in the gene Folliculin (FLCN) were first identified in BHD patients in 2002. In addition FLCN mutations have also been described in families with isolated primary spontaneous pneumothorax (PSP) and also familial clear cell renal carcinomas (FcRCC). We have established a locus-specific database based on the Leiden Open (source) Variation Database (LOVD) software. The version of the database contains 60 previously published mutations and 10 previously unpublished novel germline FLCN mutations. The mutations are comprised of deletions (44.3%), substitutions (35.7%), duplications (14.3%) and deletion/insertions (5.7%). The database is accessible online at http://www.lovd.nl/flcn.
Assuntos
Bases de Dados Genéticas , Éxons/genética , Íntrons/genética , Mutação , Proteínas Proto-Oncogênicas/genética , Proteínas Supressoras de Tumor/genética , Folículo Piloso/patologia , Humanos , Internet , Neoplasias Renais/genética , Pneumotórax/genética , Dermatopatias Genéticas/genética , SíndromeRESUMO
Birt-Hogg-Dubé (BHD) syndrome is a rare autosomal dominant condition caused by mutations in the FLCN gene and characterized by benign hair follicle tumors, pneumothorax, and renal cancer. Folliculin (FLCN), the protein product of the FLCN gene, is a poorly characterized tumor suppressor protein, currently linked to multiple cellular pathways. Autophagy maintains cellular homeostasis by removing damaged organelles and macromolecules. Although the autophagy kinase ULK1 drives autophagy, the underlying mechanisms are still being unraveled and few ULK1 substrates have been identified to date. Here, we identify that loss of FLCN moderately impairs basal autophagic flux, while re-expression of FLCN rescues autophagy. We reveal that the FLCN complex is regulated by ULK1 and elucidate 3 novel phosphorylation sites (Ser406, Ser537, and Ser542) within FLCN, which are induced by ULK1 overexpression. In addition, our findings demonstrate that FLCN interacts with a second integral component of the autophagy machinery, GABA(A) receptor-associated protein (GABARAP). The FLCN-GABARAP association is modulated by the presence of either folliculin-interacting protein (FNIP)-1 or FNIP2 and further regulated by ULK1. As observed by elevation of GABARAP, sequestome 1 (SQSTM1) and microtubule-associated protein 1 light chain 3 (MAP1LC3B) in chromophobe and clear cell tumors from a BHD patient, we found that autophagy is impaired in BHD-associated renal tumors. Consequently, this work reveals a novel facet of autophagy regulation by ULK1 and substantially contributes to our understanding of FLCN function by linking it directly to autophagy through GABARAP and ULK1.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Autofagia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Sequência de Aminoácidos , Animais , Proteínas Reguladoras de Apoptose , Proteína Homóloga à Proteína-1 Relacionada à Autofagia , Síndrome de Birt-Hogg-Dubé/metabolismo , Síndrome de Birt-Hogg-Dubé/patologia , Proteínas de Transporte/metabolismo , Humanos , Camundongos , Dados de Sequência Molecular , Fosforilação , Ligação Proteica , Proteínas Proto-Oncogênicas/química , Proteínas Proto-Oncogênicas/deficiência , Proteína Sequestossoma-1 , Proteínas Supressoras de Tumor/química , Proteínas Supressoras de Tumor/deficiênciaRESUMO
Birt-Hogg-Dubé syndrome (MIM #135150) is characterized by the development of benign skin tumours called fibrofolliculomas, pulmonary cysts that may lead to pneumothorax and a high risk of developing kidney cancer. BHD is caused by mutations affecting the highly conserved protein folliculin (FLCN), which probably has a role in intracellular transport. Most of the research effort directed towards BHD has focused on understanding how loss of FLCN causes kidney cancer. The cutaneous manifestations have received comparatively little attention. Although understandable, it is unfortunate, as the fibrofolliculomas are highly accessible and thus potentially are an excellent system for trying to understand the basic pathobiology of BHD. Also, patients can be very much burdened by the cosmetic consequences of having hundreds of facial skin tumours. Our lack of insight into what drives fibrofolliculoma growth translates into a very limited therapeutic arsenal. Thus, paying attention to fibrofolliculomas has both basic science and practical benefits. In this review, we will discuss the state of the art regarding our understanding of fibrofolliculoma pathogenesis and indicate future directions for research.
Assuntos
Síndrome de Birt-Hogg-Dubé/complicações , Dermatopatias/etiologia , Humanos , Dermatopatias/diagnóstico , Dermatopatias/terapiaRESUMO
BACKGROUND: A patient with Birt-Hogg-Dubé syndrome (BHD) presented with gross hematuria of 6 months' duration. Imaging revealed the presence of a mass in the left prostatic lobe, in addition to a previously observed renal mass. Prostate biopsy and imaging findings indicated an inflammatory etiology, and the patient was discharged. 5 months later, the patient presented once again with urinary retention. During transurethral resection of the prostate, a mass adjacent to the bladder was observed. Postoperative imaging revealed a large pelvic mass, a second mass impinging on the rectum, and extensive lymphadenopathy. The patient died 2 weeks later. INVESTIGATIONS: CT and MRI, physical examination, measurement of serum markers, urinalysis, transrectal prostate biopsy, histopathological and genetic examination of tumor specimens, postmortem immunohistochemical analysis. DIAGNOSIS: Neuroendocrine carcinoma of prostate or bladder origin. MANAGEMENT: The patient died before planned chemotherapy or radiation therapy could be implemented. More-frequent monitoring of the patient might have led to earlier diagnosis and allowed treatment to be started before widespread tumor metastasis and invasion.