CDK5RAP2 Is Required to Maintain the Germ Cell Pool during Embryonic Development

Abstract

SummaryGene products linked to microcephaly have been studied foremost for their role in brain development, while their function in the development of other organs has been largely neglected. Here, we report the critical role of Cdk5rap2 in maintaining the germ cell pool during embryonic development. We highlight that infertility in Cdk5rap2 mutant mice is secondary to a lack of spermatogenic cells in adult mice as a result of an early developmental defect in the germ cells through mitotic delay, prolonged cell cycle, and apoptosis.

Congenital microcephaly‐linked CDK5RAP2 affects eye development

“…Because Cdk5rap2 is a centrosomal protein ubiquitously expressed in various tissues (Issa et al., ; Lizarraga et al., ), we hypothesized that other organs are also likely affected. Cdk5rap2 is also affecting cell‐cycle progression (Zaqout et al., ), and therefore abnormalities during retinal progenitor cell (RPC) proliferation and differentiation are highly expected. Here we report that an/an mice exhibit eye malformations already at early developmental stages.…”

Immunofluorescence Staining of Paraffin Sections Step by Step

“…The tissues shown in this study were fixed in 4% PFA at 4 • C for a time period ranging from 10 min (P0 eyes) via 4 h (adult testes and skeletal muscles) to overnight (adult brains). We achieved the best staining result of the testicular seminiferous tubules using this fixative and after puncturing the thick capsule of the testes (Figure 5C; Zaqout et al, 2017). The testes can be alternatively fixed in a mixture of 3.7% formalin with 0.2% glutaraldehyde and 0.05% saponin (Benson and Busch, 1996) or in Bouin solution (Ellenburg et al, 2020).…”

“…After dissection, the tissues used in this study were fixed in 4% paraformaldehyde (PFA) in 0.12 M phosphate buffer (TPO 4 ) overnight (adult brains), for 4 h (adult testes and skeletal muscles), or for 10 min (P0 eyes) at 4 • C. The thick capsule of adult testes was punctured at the beginning with a needle to allow rapid penetration of the fixative (Zaqout et al, 2017). Tissues were then dehydrated in an ethanol series, cleared with xylene, and embedded in paraffin, as described previously (Canene-Adams, 2013;Sadeghipour and Babaheidarian, 2019).…”

Primary microcephaly with an unstable genome

“…CDK5RAP2 is also required for normal germ cell cycle progression, as confirmed by the labeling of heterozygous pregnant mice with two different thymidine analogs: 5-iodo-20-deoxyuridine and 5-bromo-20-deoxyuridine (BrdU) (Zaqout et al 2017). Depleted or otherwise nonfunctional CDK5RAP2 seems to result in mitotic delay, and the decreased number of neuronal progenitors observed in Cdk5rap2 an/an mice correlates with an increased mitotic index, suggesting delayed mitosis (Lizarraga et al 2010).…”

“…The implications of CDK5RAP2 deficiency, and the resultant mitotic delay and germ cells depletion, are farreaching. For example, a severe reduction in testis volume due to a massive loss of germ cells have been reported in a CDK5RAP2-deficient mouse model (Zaqout et al 2017). CDK5RAP2 also serves an important role during physiological eye development, as demonstrated by the eye malformations observed in Cdk5rap2 an/an -mutant mice.…”