MKKS Is a Centrosome-shuttling Protein Degraded by Disease-causing Mutations via CHIP-mediated Ubiquitination

Abstract

McKusick-Kaufman syndrome (MKKS) is a recessively inherited human genetic disease characterized by several developmental anomalies. Mutations in the MKKS gene also cause Bardet-Biedl syndrome (BBS), a genetically heterogeneous disorder with pleiotropic symptoms. However, little is known about how MKKS mutations lead to disease. Here, we show that disease-causing mutants of MKKS are rapidly degraded via the ubiquitin-proteasome pathway in a manner dependent on HSC70 interacting protein (CHIP), a chaperone-dependent ubiquitin ligase. Although wild-type MKKS quickly shuttles between the centrosome and cytosol in living cells, the rapidly degraded mutants often fail to localize to the centrosome. Inhibition of proteasome functions causes MKKS mutants to form insoluble structures at the centrosome. CHIP and partner chaperones, including heat-shock protein (HSP)70/heat-shock cognate 70 and HSP90, strongly recognize MKKS mutants. Modest knockdown of CHIP by RNA interference moderately inhibited the degradation of MKKS mutants. These results indicate that the MKKS mutants have an abnormal conformation and that chaperone-dependent degradation mediated by CHIP is a key feature of MKKS/BBS diseases. INTRODUCTIONMcKusick-Kaufman syndrome (MKKS) is a recessively inherited human genetic disease predominantly characterized by developmental anomalies, including vaginal atresia with hydrometrocolpos, polydactyly, and congenital heart defects (McKusick et al., 1964). The MKKS gene encodes a 570-amino acid polypeptide with weak but significant similarity to group II chaperonins . Mutations in the MKKS gene also cause Bardet-Biedl syndrome (BBS), a genetically heterogeneous disorder characterized by obesity, retinal dystrophy, polydactyly, mental retardation, renal malformation, and hypogenitalism (Beales et al., 1999), and thus MKKS is also called BBS6. The MKKS mRNA is widely expressed in various tissues, including those affected by MKKS and BBS diseases . Although more than 10 mutations in the MKKS gene have been identified in patients (Beales et al., 2001;Slavotinek et al., 2002), little is known about how they cause MKKS and BBS.Twelve of the genes that are responsible for BBS (BBS1-12) have been identified so far, and the products of several of these genes have been suggested to be involved in intraflagellar transport in cilia and intracellular trafficking in the cytosol Badano et al., 2005;Beales, 2005;Blacque and Leroux, 2006). Cargo-carrying motors play crucial roles in intraflagellar and intracellular transport systems by bidirectionally moving on microtubules, and BBS4 interacts with the dynactin complex, which mediates interactions between cargoes and the dynein motor (Kim et al., 2004). Retrograde transport of melanosomes in zebrafish is slowed by knockdown of BBS2 or BBS4-8 (Yen et al., 2006), whereas mutations in Caenorhabditis elegans BBS7 and BBS8 cause delays in intraflagellar transport driven by kinesin motors (Ou et al., 2005). The Chlamydomonas homologue of BBS5 is essential for flagellum formation (Li et al., ...

Functional analyses of variants reveal a significant role for dominant negative and common alleles in oligogenic Bardet–Biedl syndrome

“…We found 48 of 49 mutations predicted to be pathogenic from human genetic analysis to be such, whereas our interpretation of the BBS1 M390R allele as a hypomorph is consistent with the M390R knock-in mouse model (42) and our prediction of BBS11 P130S to be pathogenic is consistent with a previous report of failure of this allele to rescue melanosome transport (15). Further, our in vivo data for BBS3 and BBS6 are in agreement with two recent biochemical studies (29,30).…”

“…We selected 12 changes (BBS2 123 IV; BBS6 C517R; BBS9 A455T; BBS10 D142N; BBS12 I170V, R235M, Q386R, and S429T; MKS1 L491I; MKS3 D261N; and L437V and RPGRIP1L G1025S) common in human populations (minor allele frequency >15%) and thereby found frequently in unaffected parents and siblings of patients, sometimes in homozygosity. We also tested a 13th allele, BBS4 N165F, an induced change that does not alter the properties of the protein (see below), and four BBS6 alleles (G52D, I339V, S511A, and R518H) that behaved indistinguishably from WT in cultured cells (29); 14 of 17 changes were benign in all three in vivo assays, suggesting a specificity of >82%. In vitro assays.…”

“…In vitro assays. Phenotypes related to some of the BBS proteins have been observed in cultured cells (8)(9)(10)(29)(30)(31)(32)(33)(34). Therefore, we asked whether some of the aberrant activity in zebrafish embryos could be explained by defective behavior in mammalian cells.…”

VariantASGR1Associated with a Reduced Risk of Coronary Artery Disease

“…1F), even though both types of the mRNA transcript were highly expressed (data not shown). However, when the transfected cells were treated with a proteasome inhibitor, which blocks the degradation of truncated and misfolded proteins, 22,23 we detected the truncated protein (Fig. 1F, and Fig.…”

Unfoldomics of Human Genetic Diseases: Illustrative Examples of Ordered and Intrinsically Disordered Members of the Human Diseasome

“…McKusick-Kaufman syndrome is characterized by developmental anomalies, including vaginal atresia with hydrometrocolpos, polydactyly, and congenital heart defects. It is associated with the mutations in the MKKS gene that encodes a centrosome-shuttling protein with weak but significant similarity to group II chaperonins [107]. MowatWilson syndrome caused by the mutations in the transcriptional repressor ZFHX1B gene on the chromosome 2q22-q23 [108].…”