Human Spermatozoa as a Model for Detecting Missing Proteins in the Context of the Chromosome-Centric Human Proteome Project

Jumeau, Fanny; Com, Emmanuelle; Lane, Lydie; Duek, Paula D.; Lagarrigue, Mélanie; Lavigne, Régis; Guillot, Laetitia; Rondel, Karine; Gateau, Alain; Melaine, Nathalie; Guével, Blandine; Sergeant, Nicolas; Mitchell, Valerie A.; Pineau, Charles

doi:10.1021/acs.jproteome.5b00170

Cited by 57 publications

(85 citation statements)

References 45 publications

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“…40 Furthermore, CFAP43 was also identified by proteomic analysis in human spermatozoa. 41 Both CFAP43 and CFAP44 contain WD repeat domains, which are highly enriched in the intraflagellar transport proteins of sperm flagella. 42,43 This evidence also supports the involvement of CFAP43 and CFAP44 in vertebrate spermatogenesis.…”

Section: Semen Parametersmentioning

confidence: 99%

“…CFAP65 was previously identified in centrioles of bovine sperm and in human spermatozoa by proteomic analyses. 40,41 CFAP65 contains a MSP (major sperm protein) domain that is present in sperm-specific proteins, and its mouse ortholog has been reported to be a putative ciliary protein. 44 Furthermore, CFAP65 is expressed specifically in sexually mature male mice but is absent in juvenile mice.…”

Section: Semen Parametersmentioning

confidence: 99%

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Biallelic Mutations in CFAP43 and CFAP44 Cause Male Infertility with Multiple Morphological Abnormalities of the Sperm Flagella

Tang¹,

Wang

Li³

et al. 2017

The American Journal of Human Genetics

242

212

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Sperm motility is vital to human reproduction. Malformations of sperm flagella can cause male infertility. Men with multiple morphological abnormalities of the flagella (MMAF) have abnormal spermatozoa with absent, short, coiled, bent, and/or irregular-caliber flagella, which impair sperm motility. The known human MMAF-associated genes, such as DNAH1, only account for fewer than 45% of affected individuals. Pathogenic mechanisms in the genetically unexplained MMAF remain to be elucidated. Here, we conducted genetic analyses by using whole-exome sequencing and genome-wide comparative genomic hybridization microarrays in a multi-center cohort of 30 Han Chinese men affected by MMAF. Among them, 12 subjects could not be genetically explained by any known MMAFassociated genes. Intriguingly, we identified compound-heterozygous mutations in CFAP43 in three subjects and a homozygous frameshift mutation in CFAP44 in one subject. All of these recessive mutations were parentally inherited from heterozygous carriers but were absent in 984 individuals from three Han Chinese control populations. CFAP43 and CFAP44, encoding two cilia-and flagella-associated proteins (CFAPs), are specifically or preferentially expressed in the testis. Using CRISPR/Cas9 technology, we generated two knockout models each deficient in mouse ortholog Cfap43 or Cfap44. Notably, both Cfap43-and Cfap44-deficient male mice presented with MMAF phenotypes, whereas the corresponding female mice were fertile. Our experimental observations on human subjects and animal models strongly suggest that biallelic mutations in either CFAP43 or CFAP44 can cause sperm flagellar abnormalities and impair sperm motility. Further investigations on other CFAP-encoding genes in more genetically unexplained MMAF-affected individuals could uncover novel mechanisms underlying sperm flagellar formation.

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Section: Semen Parametersmentioning

confidence: 99%

Section: Semen Parametersmentioning

confidence: 99%

Biallelic Mutations in CFAP43 and CFAP44 Cause Male Infertility with Multiple Morphological Abnormalities of the Sperm Flagella

Tang¹,

Wang

Li³

et al. 2017

The American Journal of Human Genetics

242

212

View full text Add to dashboard Cite

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“…Some of these criteria were also used in a concomitant study 14 involving transchromosome-based data analysis on a high quality mass spectrometry data set to catalogue missing proteins in total protein extracts from isolated human spermatozoa. This analysis validated 89 missing proteins based on version 1.0 of the HPP guidelines (http://www.thehpp.org/guidelines/).…”

mentioning

confidence: 99%

Looking for Missing Proteins in the Proteome of Human Spermatozoa: An Update

et al. 2016

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The Chromosome-Centric Human Proteome Project aims to identify proteins classed as « missing » in the neXtProt knowledgebase. In this article, we present an in-depth proteomics analysis of the human sperm proteome to identify testis-enriched missing proteins. Using a range of protein extraction procedures and LC-MS/MS analysis, we detected a total of 235 proteins (PE2-PE4) for which no previous evidence of protein expression was annotated. Through a combination of LC-MS/MS and LC-PRM analysis, data mining and immunohistochemistry, we were able to confirm the expression of 206 missing proteins (PE2-4) in line with current HPP guidelines (version 2.0). Parallel Reaction Monitoring (PRM) acquisition combined with synthetic heavy labeled peptides was used to target 36 « one-hit wonder » candidates selected on the basis of prior PSM assessment. Of this subset of candidates, 24 were validated with additional predicted and specifically targeted peptides. Evidence was found for a further 16 missing proteins using immunohistochemistry on human testis sections. The expression pattern for some of these proteins was specific to the testis, and they could potentially be valuable markers with applications in fertility assessment. Strong evidence was also found for the existence of 4 proteins labeled as "uncertain" (PE5); the status of these proteins should therefore be re-examined.Our results show how the use of a range of sample preparation techniques combined with MS-based analysis, expert knowledge and complementary antibody-based techniques can produce data of interest to the community.

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“…Our laboratory recently developed a method for extracting human proteins from individual sperm ejaculates [6]. One-dimensional PAGE provides a single-step overview of sperm proteins.…”

Section: Introductionmentioning

confidence: 99%

Gel electrophoresis of human sperm: a simple method for evaluating sperm protein quality

Jumeau

Sigala

Fernandez-Gomez

et al. 2018

Basic Clin. Androl.

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BackgroundThe limitations of conventional sperm analyses have highlighted the need for additional means of evaluating sperm quality.MethodsIn a study of a cohort of 245 men with known conventional sperm parameters, one-dimensional PAGE was used to monitor protein content and quality in samples from individual ejaculates.ResultsThe sperm protein content varied markedly from sample to another, especially in the high-molecular-weight range. The intensity of the 80–110 kDa bands was correlated with progressive motility (r = 0.15, p = 0.015) and was significantly higher (p = 0.0367) in the group of men with conventional parameters above the World Health Organization’s 2010 reference values than in the group with at least one subnormal parameter (i.e. semen volume, sperm concentration, sperm count per ejaculate, progressive motility, proportion of normal forms or multiple anomaly index below the lower reference value). Using mass spectrometry, the 80–110 kDa bands were found to correspond primarily to three proteins from the flagellum’s fibrous sheath: A-kinase anchor protein 4, A-kinase anchor protein 3, and spermatogenic cell-specific type 1 hexokinase.ConclusionOne-dimensional PAGE constitutes a simple, rapid, reliable, inexpensive method for analyzing proteins associated with sperm motility in individual human ejaculates.

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Human Spermatozoa as a Model for Detecting Missing Proteins in the Context of the Chromosome-Centric Human Proteome Project

Cited by 57 publications

References 45 publications

Biallelic Mutations in CFAP43 and CFAP44 Cause Male Infertility with Multiple Morphological Abnormalities of the Sperm Flagella

Biallelic Mutations in CFAP43 and CFAP44 Cause Male Infertility with Multiple Morphological Abnormalities of the Sperm Flagella

Looking for Missing Proteins in the Proteome of Human Spermatozoa: An Update

Gel electrophoresis of human sperm: a simple method for evaluating sperm protein quality

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