BACKGROUND: The potassium channel tetramerization domain containing 13 (KCTD13) protein is a substrate-specific adapter for cullin3-based E3 ubiquitin ligase. KCTD13 is located at the 16p11.2 locus and has been associated with neurodevelopmental and neuropsychiatric disorders. Patients with copy number variants at this locus exhibit genitourinary (GU) tract anomalies.
METHODS: Equal amounts of testicular homogenate from WT and KCTD13-/- mice were subjected to IP using AR antibody. HEK293 cells were transfected as indicated and the total cell lysates were immunoprecipitated using the indicated antibodies. The resulting immunocomplexes were immunoblotted as indicated. We also used immunofluorescence and gene expression assays.
RESULTS: We found that androgen receptor (AR) was markedly down-regulated in the testis of Kctd13-deficient mice. Furthermore, reduced AR protein levels correlated with increased AR ubiquitination. Ectopic expression of KCTD13 decreased STUB1-mediated AR ubiquitination by blocking STUB1 binding to AR. KCTD13 overexpression increased CUL3-mediated AR ubiquitination but had no effect on CUL3 binding to AR. Interestingly, when equal amounts of KCTD13 and STUB1 were co-transfected, KCTD13 increased STUB1 binding to AR resulting in increased AR ubiquitination. The BTB domain of KCTD13 is required for binding both the N-terminal domain of AR and STUB1. KCTD13 ΔBTB was unable to prevent STUB1 binding to AR and subsequent STUB1-mediated AR ubiquitination. KCTD13 increased the expression of AR target gene, FOXJ1, whereas KCTD13 ΔBTB had no effect.
CONCLUSIONS: Taken together, our data reveal an interesting first step toward understanding the molecular basis of impaired AR ubiquitination seen in testicular tissues of Kctd13-deficient mice. We can further hypothesize that in Kctd13-deficient mice, the absence of KCTD13 results in increased association between AR and STUB1 leading to aberrant AR ubiquitination and degradation through the proteasomal pathway, thus connecting impaired AR ubiquitination to GU defects. Therefore, KCTD13 could be considered as a potential therapeutic target to suppress AR nuclear accumulation.
