The Expression of Cysteine-Rich Secretory Protein 2 (CRISP2) and Its Specific Regulator miR-27b in the Spermatozoa of Patients with Asthenozoospermia1

Abstract

Cysteine-rich secretory protein 2 (CRISP2) is an important sperm protein and plays roles in spermatogenesis, modulation of flagellar motility, acrosome reaction, and gamete fusion. Clinical evidence shows a reduced CRISP2 expression in spermatozoa from asthenozoospermic patients, but the molecular mechanism underlying its reduction remains unknown. Herein, we carried out a study focusing on the CRISP2 reduction and its roles in asthenozoospermia. Initially, through analyzing CRISP2 expression and methylation on CRISP2 promoter activity in sperm, we observed a decreased expression of CRISP2 protein rather than its mRNA in the ejaculated spermatozoa from asthenozoospermic patients and no methylation in the CRISP2 promoter, suggesting CRISP2 expression may be regulated in the sperm at the posttranscriptional level. Subsequently, we found that microRNA 27b (miR-27b), predicted as a candidate regulator of CRISP2 using bioinformatics, was highly expressed in the ejaculated spermatozoa from asthenozoospermic patients. Luciferase reporter assay and transfection experiments disclosed that this microRNA could target CRISP2 by specifically binding its 3' untranslated region, suppressing CRISP2 expression. Extended clinical observation further confirmed a highly expressed miR-27b and its obviously negative correlation with CRISP2 protein expression in ejaculated spermatozoa samples from asthenozoospermic patients. Finally, we conducted a retrospective follow-up study to support that either high miR-27b expression or low CRISP2 protein expression was significantly associated with low sperm progressive motility, abnormal morphology, and infertility. Thus, this study provides the first preliminary insight into the mechanism leading to the reduced CRISP2 expression in asthenozoospermia, offering a potential therapeutic target for treating male infertility or for male contraception.

Posttesticular sperm maturation, infertility, and hypercholesterolemia

“…Furthermore, in a retrospective study, high miR-27b expression or low CRISP2 protein expression was significantly associated with low sperm progressive motility, abnormal morphology, and infertility. 85 MiR-27b was earlier described as a regulator of adipocyte differentiation, as expression of miR-27 resulted in blockade of expression of PPARg and C/EBPα, the two master regulators of adipogenesis 86 and thus could represent a link between lipid metabolism, sperm epididymal maturation, and male infertility in humans.…”

MicroRNA-27a-mediated repression of cysteine-rich secretory protein 2 translation in asthenoteratozoospermic patients

“…CRISP2 is a sperm protein that influences sperm motility as well as sperm morphology. Interestingly, our previous study 36 and this study identified different upstream regulators of CRISP2 in AZ and ATZ; miR-27b was highly expressed in AZ, mainly inhibiting sperm motility by suppressing CRISP2 , whereas miR-27a was highly expressed in ATZ, mainly causing sperm malformation by repressing the same target gene. This implies the heterogeneity or specificity of miRNAs in various biological conditions.…”

“…Consistent with this, in our previous work, we found that miR-27b could target the 3’- UTR of CRISP2 and downregulate its expression in asthenozoospermia. 36 In this work, we observed that miR-27a is another specific regulator of CRISP2 and it could mediate translational repression of CRISP2 in the spermatozoa of patients with ATZ.…”

A novel role for ribonuclease 9 in suppressing sperm capacitation and acrosome reaction

“…Parameters of the sperms, including semen volume, pH, morphology, progressive motility (PR) and concentration, were classified to be normal, according to the criteria of World Health Organization (WHO, 2010). The sperms were separated by 45-90% percoll gradient centrifugation (Pharmacia, USA) 13 . Sperms were washed 3 times at 700g for 18 min and resuspended with Biggers, Whitten, and Whittingham (BWW) medium (Sigma).…”