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Chinese Journal of Medical Genetics ; (6): 807-814, 2023.
Article in Chinese | WPRIM | ID: wpr-981827


OBJECTIVE@#To explore the correlation of mitochondrial DNA (mtDNA) variants and coronary heart disease (CHD) in a Chinese pedigree and the possible molecular mechanisms.@*METHODS@#A Chinese pedigree featuring matrilineal inheritance of CHD who visited Hangzhou First People's Hospital in May 2022 was selected as the study subject. Clinical data of the proband and her affected relatives was collected. By sequencing the mtDNA of the proband and her pedigree members, candidate variants were identified through comparison with wild type mitochondrial genes. Conservative analysis among various species was conducted, and bioinformatics software was used to predict the impact of variants on the secondary structure of tRNA. Real-time PCR was carried out to determine the copy number of mtDNA, and a transmitochondrial cell line was established for analyzing the mitochondrial functions, including membrane potential and ATP level.@*RESULTS@#This pedigree had contained thirty-two members from four generations. Among ten maternal members, four had CHD, which yielded a penetrance rate of 40%. Sequence analysis of proband and her matrilineal relatives revealed the presence of a novel m.4420A>T variant and a m.10463T>C variant, both of which were highly conserved among various species. Structurally, the m.4420A>T variant had occurred at position 22 in the D-arm of tRNAMet, which disrupted the 13T-22A base-pairing, while the m.10463T>C variant was located at position 67 in the acceptor arm of tRNAArg, a position critical for steady-state level of the tRNA. Functional analysis revealed that patients with the m.4420A>T and m.10463T>C variants exhibited much fewer copy number of mtDNA and lower mitochondrial membrane potential (MMP) and ATP contents (P < 0.05), which were decreased by approximately 50.47%, 39.6% and 47.4%, respectively.@*CONCLUSION@#Mitochondrial tRNAMet 4420A>T and tRNAArg 10463T>C variants may underlay the maternally transmitted CHD in this pedigree, which had shown variation in mtDNA homogeneity, age of onset, clinical phenotype and other differences, suggesting that nuclear genes, environmental factors and mitochondrial genetic background have certain influence on the pathogenesis of CHD.

Humans , Female , Mutation , Pedigree , RNA, Transfer, Met , East Asian People , RNA, Transfer, Arg , DNA, Mitochondrial/genetics , Coronary Disease/genetics , Adenosine Triphosphate
Acta sci., Biol. sci ; 39(4): 463-467, Oct. - Dec. 2017. ilus
Article in English | LILACS | ID: biblio-877679


All living organisms need a DNA replication mechanism and it has been conserved in the three domains of life throughout evolutionary process. Primase is the enzyme responsible for synthesizing de novo RNA primers in DNA replication. Archaeo-Eukaryotic Primase (AEP) is the superfamily that typically forms a heterodimeric complex containing both a small catalytic subunit (PriS) and a large accessory noncatalytic subunit (PriL). Sulfolobus solfataricus is a model organism for research on the Genetics field. The aim of this work was to evaluate, via Bioinformatics tools, three mutations in the large subunit (PriL) of the archaeon Sulfolobus solfataricus. The aspartic acid residue in the positions (Asp) 62, (Asp) 235, (Asp) 241 have been substituted by glutamic acid (Glu). The highest positive free energy variation of the three substitutions analyzed occurred with the mutation at the (Asp) 241 site. The in silico analysis suggested that these mutations in PriL may destabilize its tridimensional structure interfering with replication mechanisms of Sulfolobus solfataricus. Moreover, it may also alter interactions with other molecules, making salt bridges, for instance.

Todos os organismos vivos necessitam de um eficiente mecanismo de replicação de DNA. Ao longo da evolução biológica foi observado que esse mecanismo é conservado nos três domínios da vida. Uma enzima importante que participa desse mecanismo é a RNA primase, a qual é responsável pela síntese de novo de iniciadores de RNA na replicação do DNA. Em Arquea-Eucariota, RNA Primase (AEP) tipicamente forma um complexo heterodimérico, que contém uma pequena subunidade catalítica (PriS) e uma subunidade maior não catalítica acessória (PriL). Sulfolobus solfataricus é um organismo modelo de Arquea para a pesquisa no campo da genética. O objetivo deste trabalho foi avaliar, por meio de ferramentas de bioinformática, três mutações pontuais na subunidade maior (PriL) de Sulfolobus solfataricus. Nas sequências mutantes, os resíduos de ácido aspártico nas posições (Asp) 62, (Asp) 235, (Asp) 241 foram substituídos por ácido glutâmico (Glu). A maior variação de energia livre positiva das três mutações analisadas ocorreu no sítio (Asp) 241. A análise in silico sugeriu que essas mutações em PriL podem desestabilizar sua estrutura tridimensional, interferindo com os mecanismos de replicação de Sulfolobus solfataricus. Além disso, podem alterar interações com outras moléculas, formando pontes salinas.

Computer Simulation , DNA Replication Timing , Sulfolobus solfataricus , Mutation , RNA, Transfer, Met
Chinese Journal of Pathology ; (12): 458-461, 2004.
Article in Chinese | WPRIM | ID: wpr-283490


<p><b>OBJECTIVE</b>To investigate variations of mtDNA in mouse tumors and to explore the relationship between mtDNA mutations and murine carcinogenesis.</p><p><b>METHODS</b>Variations of D-loop, ND3 and tRNAIle + Glu + Met gene fragments of mtDNA from six mouse tumor cell lines were analyzed by PCR-RFLP and PCR-SSCP techniques.</p><p><b>RESULTS</b>ND3 and tRNAIle + Glu + Met gene fragments of mtDNA from the tumors showed no variations at 27 endonuclease sites. The D-loop of mtDNA from Hca-F demonstrated an additional endonuclease site of Hinf I in contrast to the inbred mouse. Upon PCR-SSCP analysis, the D-loop of mtDNA was found to possess mutations in 4 of 6 tumors.</p><p><b>CONCLUSION</b>D-loop appears to be the hot spot for tumor mtDNA mutations, which may contribute to the carcinogenesis of murine tumors.</p>

Animals , Mice , Cell Line, Tumor , DNA, Mitochondrial , Genetics , DNA, Neoplasm , Genetics , Electron Transport Complex I , Mice, Inbred BALB C , Mice, Inbred C3H , Mice, Inbred C57BL , Mutation , Neoplasms, Experimental , Genetics , Pathology , Polymerase Chain Reaction , Polymorphism, Restriction Fragment Length , Polymorphism, Single-Stranded Conformational , Proteins , Genetics , RNA, Transfer, Glu , Genetics , RNA, Transfer, Ile , Genetics , RNA, Transfer, Met , Genetics