Impact of the genetic background on the reproductive phenotype of CRISP1 and CRISP4 knockout mice

PIGNATARO O; CARVAJAL G; DA ROS V; BRUKMAN NG; CUASNICÚ PS ; BATTISTONE MA; WEIGEL MUÑOZ M

Miami

Conferencia; ASA 43rd Annual Conference; 2017

American Society of Andrology

Epididymal CRISP1 and CRISP4 associate withsperm during maturation and participate in fertilization very likely throughtheir ability to regulate critical sperm Ca2+ channels. In spite of this,CRISP1 and CRISP4 knockout (KO) mice are fertile. As it has been reported thata single mutation can produce markedly different phenotypes, includingfertility, depending on the genetic background of the animals, the aim of thepresent work was to study the reproductive phenotype of CRISP1 and CRISP4 KOmice having a different genetic background. The CRISP1 and CRISP4 KO mice usedin this study had a homogeneous C57BL6 background (generated by backcrossingthe original 129SvEv/C57BL6N mice), and a mixed C57BL6/DBA background,respectively. Fertility was evaluated by mating, total motility by CASA,tyrosine phosphorylation (pTyR) by Western blot, cAMP levels by RIA,intracellular Ca2+ levels by flow cytometry, and fertilizing ability by IVF.Results revealed that males from the new CRISP1 and CRISP4 KO colonies remainedfertile but their sperm exhibited changes in several parameters compared to theoriginal colonies. Differently from CRISP1 KO sperm from the mixed geneticbackground which had clearly reduced pTyR, CRISP1 KO sperm from the newhomogeneous colony had normal pTyR but significantly reduced sperm motility.Interestingly, both reduced pTyR and motility could be restored by exposure ofsperm to dbcAMP+IBMX, suggesting a deficiency in cAMP levels in cells from thetwo genetic backgrounds. Subsequent studies confirmed the presence of lowercAMP levels in both groups, indicating that the same cAMP defect could bemanifested in two different ways depending on the geneticbackground of the animal. For CRISP4, differently from what have been observedin the two reported colonies, sperm from the new colony showed significantlylower pTyR levels, normal sperm−ZP binding, and lower fusion ability than wildtype sperm. These sperm also showed a lower increase in intracellular Ca2+during capacitation and responded differentially to Ca2+ ionophore thancontrols. Together, these observations revealed that CRISP1 and CRISP4 KOconstitute new examples of the influence of the genetic background on thereproductive phenotype. Considering the wide range of genetic backgroundsinherent to humans, these results highlight the relevance of analysing thephenotype of a particular mutation in different inbred strains to improve thediagnosis and treatment of human infertility.