Loss of mitogen-activated protein kinase kinase kinase 4 (MAP3K4) reveals a requirement for MAPK signalling in mouse sex determination

Debora Bogani, Pam Siggers, Rachel Brixey, Nick Warr, Sarah Beddow, Jessica Edwards, Debbie Williams, Dagmar Wilhelm, Peter Koopman, Richard A. Flavell, Hongbo Chi, Harry Ostrer, Sara Wells, Michael Cheeseman, Andy Greenfield

Research output: Contribution to journalArticle

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Abstract

Sex determination in mammals is controlled by the presence or absence of the Y-linked gene SRY. In the developing male (XY) gonad, sex-determining region of the Y (SRY) protein acts to up-regulate expression of the related gene, SOX9, a transcriptional regulator that in turn initiates a downstream pathway of testis development, whilst also suppressing ovary development. Despite the requirement for a number of transcription factors and secreted signalling molecules in sex determination, intracellular signalling components functioning in this process have not been defined. Here we report a role for the phylogenetically ancient mitogen-activated protein kinase (MAPK) signalling pathway in mouse sex determination. Using a forward genetic screen, we identified the recessive boygirl (byg) mutation. On the C57BL/6J background, embryos homozygous for byg exhibit consistent XY gonadal sex reversal. The byg mutation is an A to T transversion causing a premature stop codon in the gene encoding MAP3K4 (also known as MEKK4), a mitogen-activated protein kinase kinase kinase. Analysis of XY byg/byg gonads at 11.5 d post coitum reveals a growth deficit and a failure to support mesonephric cell migration, both early cellular processes normally associated with testis development. Expression analysis of mutant XY gonads at the same stage also reveals a dramatic reduction in Sox9 and, crucially, Sry at the transcript and protein levels. Moreover, we describe experiments showing the presence of activated MKK4, a direct target of MAP3K4, and activated p38 in the coelomic region of the XY gonad at 11.5 d post coitum, establishing a link between MAPK signalling in proliferating gonadal somatic cells and regulation of Sry expression. Finally, we provide evidence that haploinsufficiency for Map3k4 accounts for T-associated sex reversal (Tas). These data demonstrate that MAP3K4-dependent signalling events are required for normal expression of Sry during testis development, and create a novel entry point into the molecular and cellular mechanisms underlying sex determination in mice and disorders of sexual development in humans.

Original languageEnglish (US)
Article numbere1000196
JournalPLoS Biology
Volume7
Issue number9
DOIs
StatePublished - 2009
Externally publishedYes

Fingerprint

MAP Kinase Kinase Kinase 4
mitogen-activated protein kinase kinase kinase
Mitogen-Activated Protein Kinases
mitogen-activated protein kinase
gonads
Gonads
gender
mice
testes
sex reversal
Testis
Sex-Determining Region Y Protein
Genes
sex linkage
MAP Kinase Kinase Kinases
mutation
Gene encoding
Mammals
stop codon
sexual development

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)
  • Neuroscience(all)

Cite this

Loss of mitogen-activated protein kinase kinase kinase 4 (MAP3K4) reveals a requirement for MAPK signalling in mouse sex determination. / Bogani, Debora; Siggers, Pam; Brixey, Rachel; Warr, Nick; Beddow, Sarah; Edwards, Jessica; Williams, Debbie; Wilhelm, Dagmar; Koopman, Peter; Flavell, Richard A.; Chi, Hongbo; Ostrer, Harry; Wells, Sara; Cheeseman, Michael; Greenfield, Andy.

In: PLoS Biology, Vol. 7, No. 9, e1000196, 2009.

Research output: Contribution to journalArticle

Bogani, D, Siggers, P, Brixey, R, Warr, N, Beddow, S, Edwards, J, Williams, D, Wilhelm, D, Koopman, P, Flavell, RA, Chi, H, Ostrer, H, Wells, S, Cheeseman, M & Greenfield, A 2009, 'Loss of mitogen-activated protein kinase kinase kinase 4 (MAP3K4) reveals a requirement for MAPK signalling in mouse sex determination', PLoS Biology, vol. 7, no. 9, e1000196. https://doi.org/10.1371/journal.pbio.1000196
Bogani, Debora ; Siggers, Pam ; Brixey, Rachel ; Warr, Nick ; Beddow, Sarah ; Edwards, Jessica ; Williams, Debbie ; Wilhelm, Dagmar ; Koopman, Peter ; Flavell, Richard A. ; Chi, Hongbo ; Ostrer, Harry ; Wells, Sara ; Cheeseman, Michael ; Greenfield, Andy. / Loss of mitogen-activated protein kinase kinase kinase 4 (MAP3K4) reveals a requirement for MAPK signalling in mouse sex determination. In: PLoS Biology. 2009 ; Vol. 7, No. 9.
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abstract = "Sex determination in mammals is controlled by the presence or absence of the Y-linked gene SRY. In the developing male (XY) gonad, sex-determining region of the Y (SRY) protein acts to up-regulate expression of the related gene, SOX9, a transcriptional regulator that in turn initiates a downstream pathway of testis development, whilst also suppressing ovary development. Despite the requirement for a number of transcription factors and secreted signalling molecules in sex determination, intracellular signalling components functioning in this process have not been defined. Here we report a role for the phylogenetically ancient mitogen-activated protein kinase (MAPK) signalling pathway in mouse sex determination. Using a forward genetic screen, we identified the recessive boygirl (byg) mutation. On the C57BL/6J background, embryos homozygous for byg exhibit consistent XY gonadal sex reversal. The byg mutation is an A to T transversion causing a premature stop codon in the gene encoding MAP3K4 (also known as MEKK4), a mitogen-activated protein kinase kinase kinase. Analysis of XY byg/byg gonads at 11.5 d post coitum reveals a growth deficit and a failure to support mesonephric cell migration, both early cellular processes normally associated with testis development. Expression analysis of mutant XY gonads at the same stage also reveals a dramatic reduction in Sox9 and, crucially, Sry at the transcript and protein levels. Moreover, we describe experiments showing the presence of activated MKK4, a direct target of MAP3K4, and activated p38 in the coelomic region of the XY gonad at 11.5 d post coitum, establishing a link between MAPK signalling in proliferating gonadal somatic cells and regulation of Sry expression. Finally, we provide evidence that haploinsufficiency for Map3k4 accounts for T-associated sex reversal (Tas). These data demonstrate that MAP3K4-dependent signalling events are required for normal expression of Sry during testis development, and create a novel entry point into the molecular and cellular mechanisms underlying sex determination in mice and disorders of sexual development in humans.",
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AU - Wilhelm, Dagmar

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