Dataset Information

Novel MKRN3 Missense Mutations Associated With Central Precocious Puberty Reveal Distinct Effects on Ubiquitination.

ABSTRACT:

Context

Loss-of-function mutations in the maternally imprinted genes, MKRN3 and DLK1, are associated with central precocious puberty (CPP). Mutations in MKRN3 are the most common known genetic etiology of CPP.

Objective

This work aimed to screen patients with CPP for MKRN3 and DLK1 mutations and analyze the effects of identified mutations on protein function in vitro.

Methods

Participants included 84 unrelated children with CPP (79 girls, 5 boys) and, when available, their first-degree relatives. Five academic medical institutions participated. Sanger sequencing of MKRN3 and DLK1 5' upstream flanking and coding regions was performed on DNA extracted from peripheral blood leukocytes. Western blot analysis was performed to assess protein ubiquitination profiles.

Results

Eight heterozygous MKRN3 mutations were identified in 9 unrelated girls with CPP. Five are novel missense mutations, 2 were previously identified in patients with CPP, and 1 is a frameshift variant not previously associated with CPP. No pathogenic variants were identified in DLK1. Girls with MKRN3 mutations had an earlier age of initial pubertal signs and higher basal serum luteinizing hormone and follicle-stimulating hormone compared to girls with CPP without MRKN3 mutations. Western blot analysis revealed that compared to wild-type MKRN3, mutations within the RING finger domain reduced ubiquitination whereas the mutations outside this domain increased ubiquitination.

Conclusion

MKRN3 mutations were present in 10.7% of our CPP cohort, consistent with previous studies. The novel identified mutations in different domains of MKRN3 revealed different patterns of ubiquitination, suggesting distinct molecular mechanisms by which the loss of MRKN3 results in early pubertal onset.

SUBMITTER: Magnotto JC

PROVIDER: S-EPMC10653150 | biostudies-literature | 2023 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Novel MKRN3 Missense Mutations Associated With Central Precocious Puberty Reveal Distinct Effects on Ubiquitination.

Magnotto John C JC Mancini Alessandra A Bird Keisha K Montenegro Luciana L Tütüncüler Filiz F Pereira Sidney A SA Simas Vitoria V Garcia Leonardo L Roberts Stephanie A SA Macedo Delanie D Magnuson Melissa M Gagliardi Priscila P Mauras Nelly N Witchel Selma F SF Carroll Rona S RS Latronico Ana Claudia AC Kaiser Ursula B UB Abreu Ana Paula AP

The Journal of clinical endocrinology and metabolism 20230601 7

<h4>Context</h4>Loss-of-function mutations in the maternally imprinted genes, MKRN3 and DLK1, are associated with central precocious puberty (CPP). Mutations in MKRN3 are the most common known genetic etiology of CPP.<h4>Objective</h4>This work aimed to screen patients with CPP for MKRN3 and DLK1 mutations and analyze the effects of identified mutations on protein function in vitro.<h4>Methods</h4>Participants included 84 unrelated children with CPP (79 girls, 5 boys) and, when available, their ...[more]

PMID: 36916482

Similar Datasets

Project description:Background: Central Precocious Puberty (CPP) is clinically defined by the development of secondary sexual characteristics before the age of 8 years in girls and 9 years in boys. To date, mutations in the coding region of KISS1, KISS1R, PROKR2, DLK1, and MKRN3 genes have been reported as causative for CPP. This study investigated the presence of causative mutations in both the promoter and the 5'-UTR regions of the MKRN3 gene. Methods: Sanger DNA sequencing was used for screening the proximal promoter and 5'-UTR region of the MKRN3 gene in a group of 73 index girls with CPP. Mutations identified were cloned in luciferase reporter gene vectors and transiently transfected in GN11 cells in order to check for changes in the activity of the MKRN3 promoter. GN11 cells were previously checked for Mkrn3 expression using lentivirus mediated knock-down. In silico analysis was implemented for the detection of changes in the mRNA secondary structure of the mutated MKRN3 5'-UTR. Results: Three novel heterozygous mutations (-166, -865, -886 nt upstream to the transcription start site) located in the proximal promoter region of the MKRN3 gene were identified in six non-related girls with CPP. Four of these girls shared the -865 mutation, one the -166, and another one the -886. A 5'-UTR (+13 nt downstream to the transcription start site) novel mutation was also identified in a girl with similar clinical phenotype. Gene reporter assay evaluated the identified promoter mutations and demonstrated a significant reduction of MKRN3 promoter activity in transfected GN11 cells. In silico analysis for the mutated 5'-UTR predicted a significant change of the mRNA secondary structure. The minimum free energy (MFE) of the mutated 5'-UTR was higher when compared to the corresponding wild-type indicating less stable RNA secondary structure. Conclusion: Our findings demonstrated novel genetic alterations in the promoter and 5'-UTR regulatory regions of the MKRN3 gene. These changes add to another region to check for the etiology of CPP.

Dataset Information

Novel MKRN3 Missense Mutations Associated With Central Precocious Puberty Reveal Distinct Effects on Ubiquitination.

Context

Objective

Methods

Results

Conclusion

Publications

Novel MKRN3 Missense Mutations Associated With Central Precocious Puberty Reveal Distinct Effects on Ubiquitination.

Similar Datasets

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