A missense in HSF2BP causing primary ovarian insufficiency affects meiotic recombination by its novel interactor C19ORF57/BRME1

  1. Natalia Felipe-Medina
  2. Sandrine Caburet
  3. Fernando Sánchez-Sáez
  4. Yazmine B Condezo
  5. Dirk G de Rooij
  6. Laura Gómez-H
  7. Rodrigo Garcia-Valiente
  8. Anne Laure Todeschini
  9. Paloma Duque
  10. Manuel Adolfo Sánchez-Martin
  11. Stavit A Shalev
  12. Elena Llano
  13. Reiner A Veitia
  14. Alberto M Pendás

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Abstract

Primary Ovarian Insufficiency (POI) is a major cause of infertility, but its etiology remains poorly understood. Using whole-exome sequencing in a family with three cases of POI, we identified the candidate missense variant S167L in HSF2BP , an essential meiotic gene. Functional analysis of the HSF2BP-S167L variant in mouse showed that it behaves as a hypomorphic allele compared to a new loss-of-function (knock-out) mouse model. Hsf2bp S167L/S167L females show reduced fertility with smaller litter sizes. To obtain mechanistic insights, we identified C19ORF57/BRME1 as a strong interactor and stabilizer of HSF2BP and showed that the BRME1/HSF2BP protein complex co-immunoprecipitates with BRCA2, RAD51, RPA and PALB2. Meiocytes bearing the HSF2BP-S167L variant showed a strongly decreased staining of both HSF2BP and BRME1 at the recombination nodules and a reduced number of the foci formed by the recombinases RAD51/DMC1, thus leading to a lower frequency of crossovers. Our results provide insights into the molecular mechanism of HSF2BP-S167L in human ovarian insufficiency and sub(in)fertility.

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  1. Version published to 10.7554/elife.56996 on eLife
  2. eLife

    ###This manuscript is in revision at eLife

    The decision letter after peer review, sent to the authors on April 27, 2020, follows.

    Summary

    This paper presents a primary ovarian insufficiency (POI) case study, proband and affected family members with a unique genetic variant in HSF2BP. To examine the functional consequences of this variant, they generated a mouse mutant with the equivalent allele (S167L). Despite ostensibly normal numbers of follicles, Hsf2bp- S167L female mice showed reduced fertility in females. The authors also identify an interaction partner of HSF2BP named MIDAP and provide evidence that these two proteins function interdependently to facilitate BRCA2-mediated assembly of RAD51/DMC1 strand-exchange complexes.

    Essential Revisions

    Overall, while there are a lot of data in this paper, the quality of some experiments is questionable and lack rigorous quantification. The text is often inaccurate, sometimes not correct and requires major clarification and improvement.

    Below are essential points and important point to revise, that include new data, formatting figures, text modification and reorganization.

    1. One essential issue is to validate the conclusion that the case of human POI is caused by a variant of HSF2BP. Thus, the mouse mutant Hsf2bp167L requires additional analysis.

    2. Another major problem is order and clarity of data presentation:

    a) When presenting the phenotype of the Hsf2bp point mutant, the authors should clarify first the already published phenotype of the null mutant. They can add the data of their own null mutant as comparison.

    b) Order of data: Males and female data are not presented in a consistent way for the analysis of Hsf2bp mutant (fig3, 4 and 5): Best would be to present male and female in parallel: Fig 3 (RPA), Fig 4 (DMC1, RAD51), Fig 5 (Mlh1). RAD51 data for female is required.

    MIDAP localization in Hsf2bp mutants should be presented along with its colocalization with HSF2BP in wt.

    Analysis of SPATA22 should be moved to main figures.

    1. Removing poorly informative data: The in vitro assay fails to detect DNA binding activity of MIDAP or HSF2BP. This negative result obtained with non-purified proteins of undetermined concentration and molar ratio with the substrate do not allow to conclude on the DNA binding property of those proteins. Only part of the co-transfection analysis (Fig10) should be maintained.

    2. The localization of HSF2BP/MIDAP at DSB repair foci is not shown in this study. This should be clarified either by referring to the previous study or by doing colocalization analysis (with RPA or DMC1 or RAD51).

    3. Clarifying interpretations: Importantly the protein levels of HSF2BP and MIDAP is the mutants (Midap and Hsfp2bp) should be tested (in extracts from juvenile to avid confounding effect of cellular composition). Any coIP and western blot analysis from mouse testis to validate the interactions? And also the one with BRCA2? This would be expected given the Mass/spec data.

  3. Version published to 10.1101/2020.03.05.978007v1 on bioRxiv