DNA Binding Activity of Recombinant SRY from Normal Males and XY Females

Harley, Vincent R.; Jiang, Di; Hextall, PJ; Hawkins, Gay; Berkovitz, GD; Sockanathan, Shanthini; Lovell-Badge, Robin; Goodfellow, P N

doi:10.1126/science.1734522

Cited by 426 publications

(260 citation statements)

References 32 publications

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“…The DNA binding domain of SRY shares homology with a number of known or suspected transcription factors of the HMG box class [2]. Experiments in vitro suggest that SRY binds linear DNA in a sequence-specific manner [3,4] and causes a bend [5,6]. Mutations in the HMG domain of SRY encoded by XY gonadal dysgenesis patients affect the DNA binding or DNA bending activity of SRY in vitro which is consistent with these activities being necessary for testis development [7].…”

Section: Introductionmentioning

confidence: 58%

“…In the presence of CaM, peptides from two regions of the SRY HMG domain showed an interaction: residues 57-80 (lane 1) and a second, weakly binding region, 131-144, which gave multiple bands (lane 9). No CaM binding was detectable with the other six SRY peptides (lanes [3][4][5][6][7][8]. Binding affinity of SRY peptides 57-80 (QDRVKRPM-NAFIVWSRDQRRKMAL) to CaM was similar to that for the whole HMG box; binding of SRY peptides 131-144 (PRRKAKMLPKNCSL) was about five-fold weaker.…”

Section: Resultsmentioning

confidence: 99%

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The HMG box of SRY is a calmodulin binding domain

Harley

Lovell-Badge²,

Goodfellow

et al. 1996

FEBS Letters

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Section: Introductionmentioning

confidence: 58%

Section: Resultsmentioning

confidence: 99%

The HMG box of SRY is a calmodulin binding domain

Harley

Lovell-Badge²,

Goodfellow

et al. 1996

FEBS Letters

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“…EMSA analysis was performed using in vitro translated protein combined with 1 μl of 32 P-labeled DNA probe (~40,000 cpm) using standard protocol (Harley et al, 1992). Oligonucleotide sequence of the SRY consensus probe was 5′-GGGTTAACTAAACAATAGAATCTGGTAGA-3′; core binding sequence is underlined.…”

Section: Electromobility Shift Assay (Emsa)mentioning

confidence: 99%

“…SRY is a member of the SOX family of proteins characterized by the presence of a DNA binding domain, the High Mobility Group (HMG) (Harley et al, 1992. The ability of SRY to bind and bend DNA in vitro was established more than fifteen years ago, however its direct target genes are yet to be identified.…”

Section: Introductionmentioning

confidence: 99%

Human SRY inhibits β-catenin-mediated transcription

Bernard

Sim

Knower

et al. 2008

The International Journal of Biochemistry & Cell Biology

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In most mammals, sex is determined by the presence or absence of the SRY gene. SRY encodes a DNA binding HMG-box transcription factor which, during embryogenesis, is the initial trigger of testis differentiation from the bipotential gonad, yet its precise mode of function remains unclear. In ovarian development, R-spondin1 and Wnt4 act through the Wnt/β-catenin signaling pathway to regulate TCF-dependent expression of unknown target genes and repress testis development. Conversely, SRY may be necessary to prevent the development of ovaries by inhibiting the action of ovarian-determining genes. We hypothesize that SRY prevents Wnt/β-catenin signaling, thereby inhibiting ovarian development. In HEK293T cells, SRY repressed β-catenin-mediated TCFdependent gene activation in the presence of a specific GSK3β inhibitor or an activated β-catenin mutant, suggesting that SRY inhibits Wnt signaling at the level of β-catenin. Three SRY mutant proteins with nuclear localization defects, encoded by XY male-to-female patients, failed to inhibit β-catenin; surprisingly four SRY sex reversed mutants with defective DNA binding activity showed near wild-type SRY inhibitory activity. Moreover the potent transactivator SRY-VP16 fusion protein also showed wild-type SRY inhibitory activity. Thus SRY inhibition of β-catenin involves neither DNA binding nor transactivation functions of SRY. β-catenin and SRY interact in-vitro and SRY expression triggered β-catenin localization into specific nuclear bodies in NT2/D1 and Hela cells. We conclude that SRY inhibits β-catenin-mediated Wnt signaling by a novel nuclear function of SRY that could be important in sex determination.

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“…Gel retardation assay was carried out using three kinds of synthetic duplex DNA : HuSRY contains an AACAAAG sequence, a high affinity site for human SRY, while HuAllmut contains a CCGCGGT sequence in exchange for the AACAAAG sequence within HuSRY [19]; Mutl 1 contains an AACAAT sequence, a high affinity site for mouse Sry and Sox5 [20]. Fig.…”

Section: Sequence Specific Dna Binding Of Rainbow Trout Soxp1mentioning

confidence: 99%

A rainbow trout SRY‐type gene expressed in pituitary glands

et al. 1995

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A Sox (SRY-type HMG box) gene, designated SoxP1, was isolated from a cDNA library made from pituitaries of immature rainbow trout. Sequence analysis indicated that the cDNA had an open reading frame encoding 467 amino acid residues containing a DNA binding motif, known as the high mobility group (HMG) box. Northern blot analysis showed trout SoxP1 mRNA was detected in pituitaries and gonadal tissues, but not in liver, spleen, and heart. In pituitaries, trout SoxP1 mRNA was more abundant in immature fish than in mature fish. Gel shift retardation analysis indicated that the recombinant HMG box protein of SoxP1 produced in E. coli had a DNA binding property for an AACAAT or AACAAAG sequence. These findings suggest that the trout SoxP1 protein may play certain roles in growth or maturation in pituitaries as a transcription factor.

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DNA Binding Activity of Recombinant SRY from Normal Males and XY Females

Cited by 426 publications

References 32 publications

The HMG box of SRY is a calmodulin binding domain

The HMG box of SRY is a calmodulin binding domain

Human SRY inhibits β-catenin-mediated transcription

A rainbow trout SRY‐type gene expressed in pituitary glands

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