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Yeast model identifies ENTPD6 as a potential non-obstructive azoospermia pathogenic gene.

Wang Q, Liu C, Tang C, Guo H, Liu Y, Wang L, Zhao H, Shang Y, Wen Y, Lin Y, Zhou T, Zhou Z, Dong W, Hu Z, Guo X, Sha J, Li W - Sci Rep (2015)

Bottom Line: Recently, a genome-wide association study (GWAS) of NOA in Han Chinese men was conducted, and only a few genetic variants associated with NOA were found, which might have resulted from genetic heterogeneity.After functional screening, GDA1 (orthologous to humanENTPD6) was found to be a novel meiosis-related gene.These yeast data suggest that ENTPD6 may be a novel meiosis-associated NOA-related gene, and the yeast model provides a good approach to analyze GWAS results of NOA.

View Article: PubMed Central - PubMed

Affiliation: 1] State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China [2] University of Chinese Academy of Sciences, Beijing 100049, China.

ABSTRACT
Approximately ten percent of male infertility is caused by non-obstructive azoospermia (NOA), but the etiologies of many NOA remain elusive. Recently, a genome-wide association study (GWAS) of NOA in Han Chinese men was conducted, and only a few genetic variants associated with NOA were found, which might have resulted from genetic heterogeneity. However, those variants that lack genome-wide significance might still be essential for fertility. Functional analysis of genes surrounding these variants in Drosophila identified some spermatogenesis-essential genes. As a complementary method of Drosophila screening, SK1 background Saccharomvces cerevisiae was used as a model to screen meiosis-related genes from the NOA GWAS data in this study. After functional screening, GDA1 (orthologous to humanENTPD6) was found to be a novel meiosis-related gene. The deletion of GDA1 resulted in the failure of yeast sporulation. Further investigations showed that Gda1p was important for pre-meiotic S phase entry. Interestingly, the meiotic role of Gda1p was dependent on its guanosine diphosphatase activity, but not it's cytoplasmic, transmembrane or stem domains. These yeast data suggest that ENTPD6 may be a novel meiosis-associated NOA-related gene, and the yeast model provides a good approach to analyze GWAS results of NOA.

No MeSH data available.


Related in: MedlinePlus

N-terminal domains of Gda1p are not necessary for its role in meiosis.(a) Schematic representation of the domains of Gda1p and some mutants, including ΔN1–9 (Δ1–9aa), ΔN10–24 (Δ10–24aa), and ΔN25–58 (Δ25–58aa). CD indicates the cytoplasmic domain; TMD indicates the transmembrane domain; SR indicates the stem region; ACR1–5 indicates the conserved amino acids related to guanosine diphosphatase activity; stars indicate the glycosylation sites. (b) The function of GDA1 in meiosis was independent of its localization-terminal domains. The WT strain harboring the empty vector and the gda1Δ strains harboring either the empty vector or GDA1, ΔN1–9, ΔN10–24, ΔN25–58 under the control of its own promoter were incubated in sporulation medium for 24 hrs. Sporulation efficiency was determined by staining with DAPI. (c) Microscopic observation of the WT strain harboring the empty vector and the gda1Δ strains harboring either the empty vector or GDA1 and its variants under the control of its own promoter after sporulation induction for 24 hrs.
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f4: N-terminal domains of Gda1p are not necessary for its role in meiosis.(a) Schematic representation of the domains of Gda1p and some mutants, including ΔN1–9 (Δ1–9aa), ΔN10–24 (Δ10–24aa), and ΔN25–58 (Δ25–58aa). CD indicates the cytoplasmic domain; TMD indicates the transmembrane domain; SR indicates the stem region; ACR1–5 indicates the conserved amino acids related to guanosine diphosphatase activity; stars indicate the glycosylation sites. (b) The function of GDA1 in meiosis was independent of its localization-terminal domains. The WT strain harboring the empty vector and the gda1Δ strains harboring either the empty vector or GDA1, ΔN1–9, ΔN10–24, ΔN25–58 under the control of its own promoter were incubated in sporulation medium for 24 hrs. Sporulation efficiency was determined by staining with DAPI. (c) Microscopic observation of the WT strain harboring the empty vector and the gda1Δ strains harboring either the empty vector or GDA1 and its variants under the control of its own promoter after sporulation induction for 24 hrs.

Mentions: Gda1p is a guanosine diphosphatase that is located in the Golgi, and it is involved in the transport of GDP-mannose into the Golgi lumen by converting guanosine diphosphate (GDP) to guanosine monophosphate (GMP) after mannose is transferred to its substrate. The GDA1 deletion strain has severe defects in the N- and O-mannosylation of proteins and glycosphingolipids4142. Gda1p has a short cytoplasmic domain in its N-terminus. The next domain is a single transmembrane domain, followed by a stem region and a large catalytic luminal domain (Fig. 4a). To further study the functional role of Gda1p in sporulation, we generated three truncations: ΔN1-9 (Gda1 Δ1–9aa, in which the cytoplasmic domain is deleted), ΔN10–24 (Gda1 Δ10–24aa, in which the transmembrane domain is deleted) and ΔN25–58 (Gda1 Δ25–58aa, in which the stem region is deleted) (Fig. 4a). These were transformed into the gda1Δ strain. We found that none of the mutants affected the sporulation efficiency compared with the WT GDA1 (Fig. 4b,c). These results suggested that the function of Gda1p in meiosis is not dependent on these three domains. Additionally, in agreement with our results, it has been reported that the cytoplasmic and transmembrane domains are not necessary for the Golgi localization of Gda1p43.


Yeast model identifies ENTPD6 as a potential non-obstructive azoospermia pathogenic gene.

Wang Q, Liu C, Tang C, Guo H, Liu Y, Wang L, Zhao H, Shang Y, Wen Y, Lin Y, Zhou T, Zhou Z, Dong W, Hu Z, Guo X, Sha J, Li W - Sci Rep (2015)

N-terminal domains of Gda1p are not necessary for its role in meiosis.(a) Schematic representation of the domains of Gda1p and some mutants, including ΔN1–9 (Δ1–9aa), ΔN10–24 (Δ10–24aa), and ΔN25–58 (Δ25–58aa). CD indicates the cytoplasmic domain; TMD indicates the transmembrane domain; SR indicates the stem region; ACR1–5 indicates the conserved amino acids related to guanosine diphosphatase activity; stars indicate the glycosylation sites. (b) The function of GDA1 in meiosis was independent of its localization-terminal domains. The WT strain harboring the empty vector and the gda1Δ strains harboring either the empty vector or GDA1, ΔN1–9, ΔN10–24, ΔN25–58 under the control of its own promoter were incubated in sporulation medium for 24 hrs. Sporulation efficiency was determined by staining with DAPI. (c) Microscopic observation of the WT strain harboring the empty vector and the gda1Δ strains harboring either the empty vector or GDA1 and its variants under the control of its own promoter after sporulation induction for 24 hrs.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4495445&req=5

f4: N-terminal domains of Gda1p are not necessary for its role in meiosis.(a) Schematic representation of the domains of Gda1p and some mutants, including ΔN1–9 (Δ1–9aa), ΔN10–24 (Δ10–24aa), and ΔN25–58 (Δ25–58aa). CD indicates the cytoplasmic domain; TMD indicates the transmembrane domain; SR indicates the stem region; ACR1–5 indicates the conserved amino acids related to guanosine diphosphatase activity; stars indicate the glycosylation sites. (b) The function of GDA1 in meiosis was independent of its localization-terminal domains. The WT strain harboring the empty vector and the gda1Δ strains harboring either the empty vector or GDA1, ΔN1–9, ΔN10–24, ΔN25–58 under the control of its own promoter were incubated in sporulation medium for 24 hrs. Sporulation efficiency was determined by staining with DAPI. (c) Microscopic observation of the WT strain harboring the empty vector and the gda1Δ strains harboring either the empty vector or GDA1 and its variants under the control of its own promoter after sporulation induction for 24 hrs.
Mentions: Gda1p is a guanosine diphosphatase that is located in the Golgi, and it is involved in the transport of GDP-mannose into the Golgi lumen by converting guanosine diphosphate (GDP) to guanosine monophosphate (GMP) after mannose is transferred to its substrate. The GDA1 deletion strain has severe defects in the N- and O-mannosylation of proteins and glycosphingolipids4142. Gda1p has a short cytoplasmic domain in its N-terminus. The next domain is a single transmembrane domain, followed by a stem region and a large catalytic luminal domain (Fig. 4a). To further study the functional role of Gda1p in sporulation, we generated three truncations: ΔN1-9 (Gda1 Δ1–9aa, in which the cytoplasmic domain is deleted), ΔN10–24 (Gda1 Δ10–24aa, in which the transmembrane domain is deleted) and ΔN25–58 (Gda1 Δ25–58aa, in which the stem region is deleted) (Fig. 4a). These were transformed into the gda1Δ strain. We found that none of the mutants affected the sporulation efficiency compared with the WT GDA1 (Fig. 4b,c). These results suggested that the function of Gda1p in meiosis is not dependent on these three domains. Additionally, in agreement with our results, it has been reported that the cytoplasmic and transmembrane domains are not necessary for the Golgi localization of Gda1p43.

Bottom Line: Recently, a genome-wide association study (GWAS) of NOA in Han Chinese men was conducted, and only a few genetic variants associated with NOA were found, which might have resulted from genetic heterogeneity.After functional screening, GDA1 (orthologous to humanENTPD6) was found to be a novel meiosis-related gene.These yeast data suggest that ENTPD6 may be a novel meiosis-associated NOA-related gene, and the yeast model provides a good approach to analyze GWAS results of NOA.

View Article: PubMed Central - PubMed

Affiliation: 1] State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China [2] University of Chinese Academy of Sciences, Beijing 100049, China.

ABSTRACT
Approximately ten percent of male infertility is caused by non-obstructive azoospermia (NOA), but the etiologies of many NOA remain elusive. Recently, a genome-wide association study (GWAS) of NOA in Han Chinese men was conducted, and only a few genetic variants associated with NOA were found, which might have resulted from genetic heterogeneity. However, those variants that lack genome-wide significance might still be essential for fertility. Functional analysis of genes surrounding these variants in Drosophila identified some spermatogenesis-essential genes. As a complementary method of Drosophila screening, SK1 background Saccharomvces cerevisiae was used as a model to screen meiosis-related genes from the NOA GWAS data in this study. After functional screening, GDA1 (orthologous to humanENTPD6) was found to be a novel meiosis-related gene. The deletion of GDA1 resulted in the failure of yeast sporulation. Further investigations showed that Gda1p was important for pre-meiotic S phase entry. Interestingly, the meiotic role of Gda1p was dependent on its guanosine diphosphatase activity, but not it's cytoplasmic, transmembrane or stem domains. These yeast data suggest that ENTPD6 may be a novel meiosis-associated NOA-related gene, and the yeast model provides a good approach to analyze GWAS results of NOA.

No MeSH data available.


Related in: MedlinePlus