Role of Kctd13 in Modulating Gene Expression in Different Penile Cell Populations
Abhishek Seth, MD1, Ahmed Chahdi, PhD1, Carolina Jorgez, PhD2.
1Nemours Children's Health Orlando, orlando, FL, USA, 2Baylor College of Medicine, Houston, TX, USA.
BACKGROUND: KCTD13 is an important regulator of gonadal development as mice lacking Kctd13 (Kctd13-/-) have micropenis and small undescended testes with significantly decreased levels of androgen receptor (AR) and Sox9. KCTD13 acts as a substrate-specific adapter of an E3 ubiquitin-protein ligase complex BTB-CUL3-RBX1. Sox9 and AR activity, expression, and localization are regulated by posttranslational modifications including ubiquitination in which KCTD13 play an important role. We hypothesized that Kctd13 positively regulates penile AR and Sox9 stability and restoration of AR in Sox9 levels in the penis of Kctd13-/- mice could rescue the abnormal phenotype.
METHODS:To restore AR expression in urethral mesenchyme and Sox9 expression in the urethral epithelium, we generated double-knockout (KO) mice that lack Kctd13 and conditionally express AR in the urethral mesenchyme after cre activation with Twistcre (Kctd13-/-;AR;Twist2cre), and conditionally express SOX9 in the urethral epithelium after cre activation with Shhcre (Kctd13-/-;SOX9;Shhcre). Penis morphology was determined using microcomputed tomography (μCT) to perform 3D penile reconstructions for comparing the genitourinary phenotype of control, single-KO, and double-KO mice. Fertility was assessed by breeding male mice from each genotypic group to a wild-type (WT) female for 6 months, and then comparing litter size, number, and frequency between the groups.
RESULTS:Using μCT, we established that Kctd13-/- mice have micropenis since the average penile length in WT mice was 5.88±0.16mm and Kctd13-/- mice penises ranged from 2.90-5.54mm with 13 penises characterized as micropenis (<5.48mm) and 60% with a length below 4mm (11.5 SD). In addition, when compared to WT mice, the male urogenital mating protuberance (MUMP) length was significantly shorter in null mice. The length and base of the baculum (bone in the murine penis) were significantly shorter and narrower in null mice compared to those in WT mice. When the double-KO mouse was compared to Kctd13-/- mice, we observed an increase in the length of the mesenchyme-derived MUMP, indicating that increase in androgen and Sox9 signal rescues the micropenis phenotype of Kctd13-/- mice. The location of the urethral meatus was similar and orthotopic in location in single and double mutant, and WT penises. The infertility of the double-KO mice was improved, indicating the importance on penile development and fertility.
CONCLUSIONS:Restoration of AR and SOX9 in the penile mesoderm and epithelium, respectively, improve penile development in Kctd13-/- mice allowing us to discern the contribution from individual cell types in the penis and the role of Kctd13 in penile development.
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