Journal Article

Functional analyses of two newly identified PITX2 mutants reveal a novel molecular mechanism for Axenfeld–Rieger syndrome

Megan Priston, Kathy Kozlowski, Dan Gill, Ken Letwin, Yvonne Buys, Alex V. Levin, Mike A. Walter and Elise Héon

in Human Molecular Genetics

Volume 10, issue 16, pages 1631-1638
Published in print August 2001 | ISSN: 0964-6906
Published online August 2001 | e-ISSN: 1460-2083 | DOI: http://dx.doi.org/10.1093/hmg/10.16.1631
Functional analyses of two newly identified PITX2 mutants reveal a novel molecular mechanism for Axenfeld–Rieger syndrome

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The specific role of PITX2 in the pathogenesis of anterior segment dysgenesis has yet to be clearly defined. We provide here new insight into PITX2 pathogenesis through mutational and functional analyses. Three PITX2 mutations were found in a screen of 38 unrelated individuals affected with anterior segment anomalies (8%). All three mutations were found among the 21 individuals affected with Axenfeld–Rieger syndrome (ARS). We have identified two novel mutations, a valine→leucine (V45L) missense mutation at position 45 within the PITX2 homeodomain, and a seven amino acid duplication (7aaDup) of residues 6–12 of the homeodomain. DNA-binding studies of the two mutant PITX2 proteins demonstrated a <10-fold reduction in the DNA-binding activity of the V45L mutant, and a >100-fold reduction in activity of the 7aaDup mutant. Luciferase reporter assays showed a >200% increase in PITX2 transactivation activity of the V45L mutant, while the 7aaDup mutant was unable to transactivate at detectable levels. Our analyses of the V45L PITX2 mutant reveal that the DNA-binding domain of PITX2 can influence transactivation activity independently of DNA binding. Furthermore, our findings expand the hypothesis that the amount of residual PITX2 activity underlies the variable severity of ocular phenotypes that result from PITX2 mutation. For the first time, we present evidence that increased PITX2 activity may underlie the severe ARS ocular phenotype. We conclude that increased activity of one PITX2 allele may be as physiologically disruptive as a mutation that nullifies a PITX2 allele, with either condition resulting in ARS.

Journal Article.  5364 words.  Illustrated.

Subjects: Genetics and Genomics

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