Analysis of mutations in mitochondrial transfer RNA genes and the maternal inheritance of polycystic ovary syndrome

Abstract

Introduction: Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of reproductive age. Despite the escalating global prevalence, there is currently no definitive predisposition test available for this condition. Among the genetic causes, variations in the mitochondrial DNA (mtDNA) are increasingly recognized as a crucial contributor to the development of PCOS. However, cross-ethnic analysis of these mutations is lacking. To fill in this gap, our objective is to identify new maternal genetic risk factors associated with PCOS by investigating the mitochondrial transfer RNA (mt-tRNA) genes in PCOS patients from Pakistan and to compare these mutations to those in patients from other ethnic groups.

Methods: DNA was extracted from saliva samples of patients. Primers were designed for the amplification of all of 22 mt-tRNA genes, and PCR was employed under defined conditions. Subsequently, Sanger sequencing was employed to decipher the sequences of mt-tRNA genes. Following sequencing, mt-tRNA genes underwent mutation analysis. Finally, we utilized MitoTIP (Mitochondrial tRNA Informatics Predictor) to identify variations in mt-tRNA genes.

Results: In a cohort of 64 Pakistani patients with PCOS, our analysis unveiled eight variants in five mt-tRNA genes including MT-TH, MT-TL2, MT-TS1, MT-TS2, and MT-TT genes. All of these variants have not been previously reported in PCOS except one we have recently identified in a Pakistani patient with PCOS. Interestingly, most of these mt-tRNA genes carry variants found in patients with PCOS across distinct ethnic groups. Furthermore, these mutations occurred in highly conserved nucleotides of tRNA, essential for ensuring the stability and biochemical functionality of mt-tRNA. Finally, the pathogenic potential of these variations was assessed by in silico analysis. The pathogenicity prediction of these variants suggests their potential impact on mitochondrial dysfunction that was responsible for the clinical phenotypes of PCOS.

Conclusion: Our study identified novel variations in mt-tRNA genes in Pakistani women with PCOS. To our knowledge, this is the first report comparing mutations of mt-tRNA genes in PCOS patients across different ethnic groups. Our data revealed common mt-tRNA genes carrying PCOS-associated mutations that may be specific to certain ethnic populations. Together, our work provides new insights into the role of mt-tRNA genes in mitochondrial dysfunction underlying the pathophysiology of PCOS, highlighting mt-tRNA mutations as potential factors for future predisposition tests and more effective therapies for this globally prevalent condition.

Keywords: PCOS; genome sequence analysis; mitochondrial tRNA genes; mutations; pathogenicity.

Figure 1

Sequence alignment and structural prediction of MT-TL2 gene. (A) Sequence alignment of MT-TL2 gene (12266-12336 nt) from maternally inherited PCOS-positive patients with rCRS Accession No. NC-012920.1 exhibiting A>G mutation at position 12307. (B) Sequence alignment of MT-TL2 gene (12266-12336 nt) from maternally inherited PCOS-positive patients with rCRS Accession No. NC-012920.1 highlighting A>G mutation at position 12307. (C) Predicted structure of normal MT-TL2 gene. (D) Predicted structure of MT-TL2 gene carrying 12267 C>T variant. (E) Predicted structure of MT-TL2 gene carrying 12308 A>G variant. (F) Predicted structure of normal MT-TL2 gene. (G) Predicted structure of MT-TL2 gene carrying 12307 A>G and 12308 A>G variant.

Figure 2

Sequence alignment and structural prediction of MT-TS1 gene. (A) Sequence alignment of MT-TS1 gene (7446-7514 nt) from maternally inherited PCOS-positive patients with rCRS Accession No. NC-012920.1 exhibiting A>G mutation at position 7490. (B) Sequence alignment of MT-TS1 gene (7446-7514 nt) from maternally inherited PCOS-positive patients with rCRS Accession No. NC-012920.1 highlighting A>G mutation at position 7490. (C) Predicted structure of normal MT-TS1 gene. (D) Predicted structure of MT-TS1 gene carrying 7490 A>G variant.

Figure 3

Sequence alignment and structural prediction of MT-TT gene. (A) Sequence alignment of MT-TT gene (15888-15953 nt) from maternally inherited PCOS-positive patients with rCRS Accession No. NC-012920.1 exhibiting G>A mutation at position 15928. (B) Sequence alignment of MT-TT gene (15888-15953 nt) from maternally inherited PCOS-positive patients with rCRS Accession No. NC-012920.1 highlighting G>A mutation at position 15928. (C) Predicted structure of normal MT-TT gene. (D) Predicted structure of MT-TT gene carrying 15928 G>A variant.

Figure 4

The family pedigree of family No. 5 (A) and No. 4 (B) based on the information provided.