SUMMARY
SPOUT1/CENP-32 encodes a putative SPOUT RNA methyltransferase previously identified as a mitotic chromosome associated protein. SPOUT1/CENP-32 depletion leads to centrosome detachment from the spindle poles and chromosome misalignment. Aided by gene matching platforms, we identified 24 individuals with neurodevelopmental delays from 18 families with bi-allelic variants in SPOUT1/CENP-32 detected by exome/genome sequencing. Zebrafish spout1/cenp-32 mutants showed reduction in larval head size with concomitant apoptosis likely associated with altered cell cycle progression. In vivo complementation assays in zebrafish indicated that SPOUT1/CENP-32 missense variants identified in humans are pathogenic. Crystal structure analysis of SPOUT1/CENP-32 revealed that most disease-associated missense variants mapped to the catalytic domain. Additionally, SPOUT1/CENP-32 recurrent missense variants had reduced methyltransferase activity in vitro and compromised centrosome tethering to the spindle poles in human cells. Thus, SPOUT1/CENP-32 pathogenic variants cause an autosomal recessive neurodevelopmental disorder: SpADMiSS (SPOUT1 Associated Development delay Microcephaly Seizures Short stature) underpinned by mitotic spindle organization defects and consequent chromosome segregation errors.
Competing Interest Statement
BCM and Miraca Holdings have formed a joint venture with shared ownership and governance of Baylor Genetics (BG), which performs clinical microarray analysis (CMA), clinical ES (cES), and clinical biochemical studies. JRL serves on the Scientific Advisory Board of the BG. The Department of Molecular and Human Genetics at Baylor College of Medicine receives revenue from clinical genetic testing conducted at BG Laboratories. JRL has stock ownership in 23andMe, is a paid consultant for Genomics International, and is a coinventor on multiple United States and European patents related to molecular diagnostics for inherited neuropathies, eye diseases, genomic disorders and bacterial genomic fingerprinting. DP provides consulting service for Ionis Pharmaceuticals. The other authors declare no competing interests.
Funding Statement
This work was supported by a grant from the US National Institute of Health (NIH) (R01MH106826) to EED, in part by US NIH (NS105078) to JRL and US NIH (UM1 HG011758) to JRL and JEP. SK is funded by an International Research Support Initiative Program (IRSIP) fellowship from the Higher Education Commission of Pakistan. EED is the Ann Marie and Francis Klocke, MD Research Scholar. IS and WCE are funded by Wellcome Principal Research Fellowship to WCE (grant no. 107022). We thank Natalia Kochanova and Shaun Webb for their help with the statistical analyses of the in vitro U2OS rescue experiments and we also thank Elizabeth Blackburn for helpful discussions on the Methyltransferase assays. MAA is funded by a grant from the Medical Research Council (MRC, United Kingdom; MR/X001245/1) to AAJ. AAJ and his team are co-funded by the European Union (ERC Advanced Grant, CHROMSEG, 101054950). Views and opinions expressed are, however, those of the authors only and do not necessarily reflect those of the European Union or the European Research Council. Neither the European Union nor the granting authority can be held responsible for them. DM was supported by a medical genetics research fellowship program through the US NIH (T32 GM007526-42). JEP was supported by NHGRI K08 HG008986. TM was supported by the Uehara Memorial Foundation. DP is supported by Doris Duke Charitable Foundation with grant# 2023-0235, and NINDS NS125126-01A1. SNW was supported in part by K08NS119567.
Author Declarations
I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.
Yes
The details of the IRB/oversight body that provided approval or exemption for the research described are given below:
Informed consent was obtained from all families Clinical evaluation and genetic analyses were approved by respective institutional ethics committees from the following participating centers Columbia University Medical Center (IRB# AAAR1159) Childrens Hospital of Philadelphia (IRB #16-013231) Baylor College of Medicine (IRB# H-29697) Nationwide Childrens Hospital (IRB# 11-00215)
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Data Availability
All data produced in the present study are available upon reasonable request to the authors