MGAT1 and Complex N-Glycans Regulate ERK Signaling During Spermatogenesis

Abstract

Mechanisms that regulate spermatogenesis in mice are important to define as they often apply to fertility in man. We previously showed that conditional deletion of the mouse Mgat1 gene (Mgat1 cKO) in spermatogonia causes a germ-cell autonomous defect leading to infertility. MGAT1 is the N-acetylglucosaminyltransferase (GlcNAcT-I) that initiates the synthesis of complex N-glycans. Mechanistic bases of MGAT1 loss were investigated in germ cells from 22- and 23-day males, before any changes in germ cell morphology were apparent. Gene expression changes induced by deletion of Mgat1 were determined using the Affymetrix gene chip Mouse Mogene 2.0 ST array, and relationships were investigated by bioinformatics including Gene Ontology (GO), Ingenuity Pathway Analysis (IPA), and Gene Set Enrichment Analysis (GSEA). The loss of complex N-glycans promoted the premature up-regulation of genes normally expressed later in spermatogenesis and spermiogenesis, and IPA and GSEA implicated ERK signaling. EGFR and PDGFRA transcripts and ERK1/2 signaling were reduced in 22-day Mgat1 cKO germ cells. Basigin, a germ cell target of MGAT1, activated ERK1/2 in CHO cells, but not in a Lec1 CHO mutant that lacks MGAT1 and complex N-glycans. Thus, MGAT1 is required to regulate ERK1/2 signaling during spermatogenesis, potentially via different mechanisms.

The dynamics of gene expression during and post meiosis sets the sperm agenda

“…Katnal1 ( Katanin p60 subunit A‐like 1) gene is required for germ cell maturation in testis and its deficiency causes temporary male sterility (Sarma et al, ), while the Kif3a gene is involved in nuclear shaping and tail formation (Ma, Wang, & Yang, ). The Mgat1 gene encodes the enzyme N‐acetylglucosaminyltransferase that regulates ERK signaling (Biswas, Batista, Sundaram, & Stanley, ), which is indispensable for spermatogenesis and Mgat1 deletion in spermatogonia did not produce any sperm (Batista, Lu, Williams, & Stanley, ). Likewise, spink2 deficient mouse have increased serine protease activity, disrupted spermatogenesis, and germ cell apoptosis (B. Lee et al, ), while the spink2 knockout homozygous male mice are azoospermic (Kherraf et al, ).…”

“…Katnal1 (Katanin p60 subunit A-like 1) gene is required for germ cell maturation in testis and its deficiency causes temporary male sterility (Sarma et al, 2017), while the Kif3a gene is involved in nuclear shaping and tail formation (Ma, Wang, & Yang, 2017). The Mgat1 gene encodes the enzyme N-acetylglucosaminyltransferase that regulates ERK signaling (Biswas, Batista, Sundaram, & Stanley, 2018), which is indispensable for spermatogenesis and Mgat1 deletion in spermatogonia did not produce any sperm (Batista, Lu, Williams, & Stanley, 2012 azoospermic (Kherraf et al, 2017). The Slc16a7 (monocarboxylate transporter-2) that transports lactate from Sertoli to germ cells was previously reported predominantly in spermatocytes and spermatids (Yadav et al, 2018) and genetic variations in MCT-2 have clinical relevance in male infertility (J.…”

The Golgi Glycoprotein MGAT4D is an Intrinsic Protector of Testicular Germ Cells From Mild Heat Stress

“…We also investigated testis lysates and testis sections for an increase in complex N-glycan levels. Basigin (CD147, BSG) carries complex and some oligomannosyl N-glycans in testis (Batista et al, 2012;Biswas et al, 2018). Thus, loss of an MGAT1 inhibitor might increase the level of complex N-glycans on BSG.…”

“…This transgene has previously been shown to inhibit MGAT1 in transfected cells (Huang et al, 2015;Huang & Stanley, 2010). The Stra8 (Stimulated By Retinoic Acid 8) promoter was used to express the transgene in spermatogonia (Batista et al, 2012;Biswas et al, 2018;Sadate-Ngatchou et al, 2008), the Ldhc (Lactate Dehydrogenase C) promoter was used to express in spermatocytes (S. Li, Zhou, Doglio, & Goldberg, 1998;Tang, Kung, & Goldberg, 2008), and the Prm1 (Protamine 1) promoter was used to express in spermatids (O'Gorman, Dagenais, Qian, & Marchuk, 1997) ( Figure 4A). The transgenic mouse strains were named Stra8-Mgat4d-L-Myc, Ldhc-Mgat4d-L-Myc and Prm1-Mgat4d-L-Myc, respectively.…”

“…Characterization of the interactions of MGAT4D in the Golgi using a fluorescence resonance energy transfer (FRET) assay showed that it interacts with MGAT1 but not MGAT2, MGAT3, MGAT4B or MGAT5 (Huang et al, 2015). Since knockout of Mgat1 in spermatogonia disrupts spermatogenesis and results in infertility (Batista, Lu, Williams, & Stanley, 2012;Biswas, Batista, Sundaram, & Stanley, 2018), deletion or overexpression of Mgat4d in germ cells were both expected to have effects on spermatogenesis. In this paper, we show that unexpectedly, deletion of Mgat4d globally, or specifically in spermatogonia, or mis-expression of Mgat4d in spermatogonia, spermatocytes or spermatids, do not appear to alter spermatogenesis in young or aged mice, and do not affect fertility.…”