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Feminizing chicks: a model for avian sex determination based on titration of Hint enzyme activity and the predicted structure of an Asw-Hint heterodimer.

Pace HC, Brenner C - Genome Biol. (2003)

Bottom Line: We consider whether positive cooperativity could explain how Hint heterodimerization with an inert enzyme might reduce specific activity by more than 50% and provide data sufficient to reject this model.Instead, we hypothesize that Asw carries a signal for mislocalization and/or proteolysis, and/or dominantly suppresses the remaining Hint active site to function as a dominant negative.Molecular modeling suggests that Asw and Hint can heterodimerize and that Gln 127, an Asw-specific alteration for Trp123, dominantly interferes with the Hint active site.

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Affiliation: Structural Biology and Bioinformatics Program, Kimmel Cancer Center, Philadelphia, PA 19107, USA.

ABSTRACT

Background: In birds and some lizards, females are heterogametic with a ZW karyotype, while males are ZZ homogametes. The molecular basis for sexual differentiation in birds is unknown: arguments exist for doses of Z masculinizing chicks and for W information feminizing. ASW was identified as a tandemly repeated gene conserved on avian W chromosomes that is expressed in early female development and appears to be an inactive form of avian Z-encoded HINT. Hint is a dimeric enzyme that hydrolyzes AMP linked to lysine, whose enzyme activity is required for regulation of the Cdk7 homologous Kin28 kinase in yeast. Of 16 residues most conserved across all life forms for AMP interactions, 15 are sexually dimorphic in birds, that is, altered in the female-specific Asw protein. Genomic and expression data suggest that Asw may feminize chicks, dominantly interfering with Hint function by heterodimerization.

Results: We consider whether positive cooperativity could explain how Hint heterodimerization with an inert enzyme might reduce specific activity by more than 50% and provide data sufficient to reject this model. Instead, we hypothesize that Asw carries a signal for mislocalization and/or proteolysis, and/or dominantly suppresses the remaining Hint active site to function as a dominant negative.

Conclusions: Molecular modeling suggests that Asw and Hint can heterodimerize and that Gln 127, an Asw-specific alteration for Trp123, dominantly interferes with the Hint active site. An extra dose of HINT in ZZW chicks, and thus more Hint homodimer, may partially overcome the feminizing influence of ASW and lead to the observed intersexual characteristics of ZZW triploids.

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Hint hydrolases are not cooperative for AMP-NH2 hydrolysis. Substrate concentration-dependent hydrolytic rates (per monomer) for rabbit Hint (filled circles) and yeast Hnt1 (open triangles) indicate that Hint hydrolases, although dimeric, have a single Km for substrate. Reproduced with permission from [30].
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Figure 2: Hint hydrolases are not cooperative for AMP-NH2 hydrolysis. Substrate concentration-dependent hydrolytic rates (per monomer) for rabbit Hint (filled circles) and yeast Hnt1 (open triangles) indicate that Hint hydrolases, although dimeric, have a single Km for substrate. Reproduced with permission from [30].

Mentions: One might expect a Hint-Asw heterodimer to have substantially less than 50% of the activity of a Hint homodimer if Hint homodimers showed cooperativity with respect to substrate binding and/or hydrolysis. For example, if the first nucleotide substrate were to bind weakly but the presence of the first bound substrate promoted efficient binding of a second substrate, then a heterodimer containing one functional and one nonfunctional active site would retain only one low-affinity binding site and be severely defective. This mechanism can be largely excluded, however, because sigmoidal substrate saturation kinetics were not observed with AMP-NH2 [30]. The data for both rabbit and yeast Hint enzymes indicate that the dimer's two crystallographically defined nucleotide-binding sites [25] bind nucleotide independently and fit well to a single submicromolar Km for each enzyme (Figure 2) [30].


Feminizing chicks: a model for avian sex determination based on titration of Hint enzyme activity and the predicted structure of an Asw-Hint heterodimer.

Pace HC, Brenner C - Genome Biol. (2003)

Hint hydrolases are not cooperative for AMP-NH2 hydrolysis. Substrate concentration-dependent hydrolytic rates (per monomer) for rabbit Hint (filled circles) and yeast Hnt1 (open triangles) indicate that Hint hydrolases, although dimeric, have a single Km for substrate. Reproduced with permission from [30].
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC153458&req=5

Figure 2: Hint hydrolases are not cooperative for AMP-NH2 hydrolysis. Substrate concentration-dependent hydrolytic rates (per monomer) for rabbit Hint (filled circles) and yeast Hnt1 (open triangles) indicate that Hint hydrolases, although dimeric, have a single Km for substrate. Reproduced with permission from [30].
Mentions: One might expect a Hint-Asw heterodimer to have substantially less than 50% of the activity of a Hint homodimer if Hint homodimers showed cooperativity with respect to substrate binding and/or hydrolysis. For example, if the first nucleotide substrate were to bind weakly but the presence of the first bound substrate promoted efficient binding of a second substrate, then a heterodimer containing one functional and one nonfunctional active site would retain only one low-affinity binding site and be severely defective. This mechanism can be largely excluded, however, because sigmoidal substrate saturation kinetics were not observed with AMP-NH2 [30]. The data for both rabbit and yeast Hint enzymes indicate that the dimer's two crystallographically defined nucleotide-binding sites [25] bind nucleotide independently and fit well to a single submicromolar Km for each enzyme (Figure 2) [30].

Bottom Line: We consider whether positive cooperativity could explain how Hint heterodimerization with an inert enzyme might reduce specific activity by more than 50% and provide data sufficient to reject this model.Instead, we hypothesize that Asw carries a signal for mislocalization and/or proteolysis, and/or dominantly suppresses the remaining Hint active site to function as a dominant negative.Molecular modeling suggests that Asw and Hint can heterodimerize and that Gln 127, an Asw-specific alteration for Trp123, dominantly interferes with the Hint active site.

View Article: PubMed Central - HTML - PubMed

Affiliation: Structural Biology and Bioinformatics Program, Kimmel Cancer Center, Philadelphia, PA 19107, USA.

ABSTRACT

Background: In birds and some lizards, females are heterogametic with a ZW karyotype, while males are ZZ homogametes. The molecular basis for sexual differentiation in birds is unknown: arguments exist for doses of Z masculinizing chicks and for W information feminizing. ASW was identified as a tandemly repeated gene conserved on avian W chromosomes that is expressed in early female development and appears to be an inactive form of avian Z-encoded HINT. Hint is a dimeric enzyme that hydrolyzes AMP linked to lysine, whose enzyme activity is required for regulation of the Cdk7 homologous Kin28 kinase in yeast. Of 16 residues most conserved across all life forms for AMP interactions, 15 are sexually dimorphic in birds, that is, altered in the female-specific Asw protein. Genomic and expression data suggest that Asw may feminize chicks, dominantly interfering with Hint function by heterodimerization.

Results: We consider whether positive cooperativity could explain how Hint heterodimerization with an inert enzyme might reduce specific activity by more than 50% and provide data sufficient to reject this model. Instead, we hypothesize that Asw carries a signal for mislocalization and/or proteolysis, and/or dominantly suppresses the remaining Hint active site to function as a dominant negative.

Conclusions: Molecular modeling suggests that Asw and Hint can heterodimerize and that Gln 127, an Asw-specific alteration for Trp123, dominantly interferes with the Hint active site. An extra dose of HINT in ZZW chicks, and thus more Hint homodimer, may partially overcome the feminizing influence of ASW and lead to the observed intersexual characteristics of ZZW triploids.

Show MeSH
Related in: MedlinePlus