High-throughput splicing assays identify missense and silent splice-disruptive POU1F1 variants underlying pituitary hormone deficiency

Peter Gergics; Cathy Smith; Hironori Bando; Alexander A L Jorge; Denise Rockstroh-Lippold; Sebastian A Vishnopolska; Frederic Castinetti; Mariam Maksutova; Luciani Renata Silveira Carvalho; Julia Hoppmann; Julián Martínez Mayer; Frédérique Albarel; Debora Braslavsky; Ana Keselman; Ignacio Bergadá; Marcelo A Martí; Alexandru Saveanu; Anne Barlier; Rami Abou Jamra; Michael H Guo; Andrew Dauber; Marilena Nakaguma; Berenice B Mendonca; Sajini N Jayakody; A Bilge Ozel; Qing Fang; Qianyi Ma; Jun Z Li; Thierry Brue; María Ines Pérez Millán; Ivo J P Arnhold; Roland Pfaeffle; Jacob O Kitzman; Sally A Camper

doi:10.1016/j.ajhg.2021.06.013

High-throughput splicing assays identify missense and silent splice-disruptive POU1F1 variants underlying pituitary hormone deficiency

Am J Hum Genet. 2021 Aug 5;108(8):1526-1539. doi: 10.1016/j.ajhg.2021.06.013. Epub 2021 Jul 15.

Authors

Peter Gergics¹, Cathy Smith², Hironori Bando¹, Alexander A L Jorge³, Denise Rockstroh-Lippold⁴, Sebastian A Vishnopolska⁵, Frederic Castinetti⁶, Mariam Maksutova¹, Luciani Renata Silveira Carvalho⁷, Julia Hoppmann⁴, Julián Martínez Mayer⁵, Frédérique Albarel⁶, Debora Braslavsky⁸, Ana Keselman⁸, Ignacio Bergadá⁸, Marcelo A Martí⁹, Alexandru Saveanu¹⁰, Anne Barlier¹⁰, Rami Abou Jamra¹¹, Michael H Guo¹², Andrew Dauber¹³, Marilena Nakaguma⁷, Berenice B Mendonca⁷, Sajini N Jayakody¹, A Bilge Ozel¹, Qing Fang¹, Qianyi Ma¹, Jun Z Li¹, Thierry Brue⁶, María Ines Pérez Millán⁵, Ivo J P Arnhold⁷, Roland Pfaeffle¹⁴, Jacob O Kitzman¹⁵, Sally A Camper¹⁶

Affiliations

¹ Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109-5618, USA.
² Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109-5618, USA; Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, MI 48109-2218, USA.
³ Genetic Endocrinology Unit (LIM25), Division of Endocrinology, Hospital das Clinicas da Faculdade de Medicina da Universidade de São Paulo, São Paulo 01246-903, Brazil.
⁴ Department of Women's and Child Health, Division of Pediatric Endocrinology, University Hospital Leipzig, Leipzig 04103, Germany.
⁵ Instituto de Biociencias, Biotecnología y Biología Traslacional, Departamento de Fisiología, Biología Molecular y Celular, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad de Buenos Aires, CABA CE1428EHA, Argentina.
⁶ Aix Marseille University, AP-HM, INSERM, Marseille Medical Genetics, Marmara Institute, La Conception Hospital, Department of Endocrinology, Marseille 13005, France.
⁷ Developmental Endocrinology Unit, Laboratory of Hormones and Molecular Genetics LIM/42, Division of Endocrinology, Hospital das Clinicas da Faculdade de Medicina da Universidade de São Paulo, São Paulo 05403-900, Brazil.
⁸ Centro de Investigaciones Endocrinológicas "Dr. César Bergadá," FEI - CONICET - División de Endocrinología, Hospital de Niños Ricardo Gutiérrez, Ciudad de Buenos Aires, CABA CE1428EHA, Argentina.
⁹ Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires e Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales CONICET, Pabellòn 2 de Ciudad Universitaria, Ciudad de Buenos Aires, CABA C1428EHA, Argentina.
¹⁰ Aix Marseille University, AP-HM, INSERM, Marseille Medical Genetics, Marmara Institute, La Conception Hospital, Laboratory of Molecular Biology, Marseille 13385, France.
¹¹ Institute of Human Genetics, University of Leipzig Medical Center, Leipzig 04103, Germany.
¹² Division of Endocrinology, Boston Children's Hospital and Department of Genetics, Harvard Medical School, Boston, MA 02115, USA.
¹³ Cincinnati Center for Growth Disorders, Division of Endocrinology, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA.
¹⁴ Department of Women's and Child Health, Division of Pediatric Endocrinology, University Hospital Leipzig, Leipzig 04103, Germany; Institute of Human Genetics, University of Leipzig Medical Center, Leipzig 04103, Germany.
¹⁵ Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109-5618, USA; Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, MI 48109-2218, USA. Electronic address: [email protected].
¹⁶ Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109-5618, USA. Electronic address: [email protected].

Abstract

Pituitary hormone deficiency occurs in ∼1:4,000 live births. Approximately 3% of the cases are due to mutations in the alpha isoform of POU1F1, a pituitary-specific transcriptional activator. We found four separate heterozygous missense variants in unrelated individuals with hypopituitarism that were predicted to affect a minor isoform, POU1F1 beta, which can act as a transcriptional repressor. These variants retain repressor activity, but they shift splicing to favor the expression of the beta isoform, resulting in dominant-negative loss of function. Using a high-throughput splicing reporter assay, we tested 1,070 single-nucleotide variants in POU1F1. We identified 96 splice-disruptive variants, including 14 synonymous variants. In separate cohorts, we found two additional synonymous variants nominated by this screen that co-segregate with hypopituitarism. This study underlines the importance of evaluating the impact of variants on splicing and provides a catalog for interpretation of variants of unknown significance in POU1F1.

Keywords: PIT-1; alternative splicing; growth hormone deficiency; hypopituitarism; massively parallel splicing assay; multiplexed assays of variant effects; transcriptional regulation; variants of uncertain significance.

Publication types

Case Reports
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Adolescent
Adult
Child
Child, Preschool
High-Throughput Screening Assays / methods*
Humans
Hypopituitarism / etiology
Hypopituitarism / metabolism
Hypopituitarism / pathology*
Male
Mutation*
Pedigree
Pituitary Hormones / deficiency*
RNA Splicing / genetics*
Transcription Factor Pit-1 / genetics*

Substances

POU1F1 protein, human
Pituitary Hormones
Transcription Factor Pit-1

Abstract

Publication types

MeSH terms

Substances

Grants and funding