Dysregulation of intracellular pH is a cause of impaired capacitation in Slc22a14-deficient mice

in Reproduction
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  • 1 Department of Applied Life Sciences, Faculty of Agriculture, Shizuoka University, Shizuoka, Japan
  • | 2 Department of Agriculture, Graduate School of Integrated Science and Technology, Shizuoka University, Shizuoka, Japan
  • | 3 Department of Reproductive and Perinatal Medicine, Hamamatsu University School of Medicine, Shizuoka, Japan
  • | 4 Tawara IVF Clinic, Shizuoka, Japan
  • | 5 Department of Virology and Parasitology, Hamamatsu University School of Medicine, Hamamatsu, Shizuoka, Japan
  • | 6 College of Agriculture, Academic Institute, Shizuoka University, Shizuoka, Japan

Correspondence should be addressed to K Yogo; Email: yogo.keiichiro@shizuoka.ac.jp
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Solute carrier 22a member 14 (SLC22A14) plays a critical role in male infertility in mice. We previously revealed that one of the causes of infertility is impaired capacitation. However, the molecular mechanism remained unclear. Here, we show that the influx of HCO3 , a trigger of capacitation, is impaired and intracellular pH (pHi) is decreased in the sperm of Slc22a14 knockout (KO) mice. While intracellular cAMP concentration did not increase during capacitation in Slc22a14 KO spermatozoa, HCO3 -dependent soluble adenylate cyclase activity was normal, and the addition of 8-bromo cAMP rescued the decreased protein tyrosine phosphorylation. In addition, the pHi of Slc22a14 KO sperm was lower than that of WT sperm and did not increase after the addition of HCO3 . Although its relationship to the regulation of pHi is unknown, transmembrane protein 225, a possible protein phosphatase inhibitor, was found to be decreased in Slc22a14 KO sperm. The decreased in vitro fertilization rate of Slc22a14 KO sperm was partially rescued by an increase in the pHi and the addition of 8-bromo cAMP. These results suggest that SLC22A14 is involved in capacitation through the regulation of HCO3 transport and pHi.

Supplementary Materials

    • Supplementary Table 1 Primer pairs and PCR condition used in semi-quantitative RT-PCR
    • Figure S1. Expression of HCO3– and H+ transporter in Slc22a14 KO mouse testis A) Expression of indicated transporters in testes were analyzed by semi-quantitative PCR. The numbers above the photo indicate the PCR cycles. The band indicated with an asterisk in Slc9a10 may be a splicing isoform. Data representative of two independent experiments are shown. B) The western blotting analysis of SLC26A3 and SLC26A6 in WT and Slc22a14 KO sperm. Data representative of two independent experiments are shown.
    • Figure S2. iTRAQ analysis using total cell lysate of sperm A) The abundance of protein in WT and KO sperm are plotted. Almost of all protein shows similar expression level. B) Top 10 protein list downregulated in Slc22a14 KO sperm.
    • Figure S3. Effect of trimethylamine on pHi of sperm BCECF–loaded WT and KO sperm were incubated in TYH-HEPES medium supplemented with 25.07 mM NaHCO3, 4 mg/ml BSA, and the indicated concentration of trimethylamine (TMA) for 30 min, and pHi was measured using a spectrophotometer.

 

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