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In this report, we investigated the molecular genetic mechanism underlying the deafness-associated mitochondrial tRNA(His) 12201T>C mutation. The destabilization of a highly conserved base-pairing (5A-68U) by the m.12201T>C mutati...
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In this report, we investigated the molecular genetic mechanism underlying the deafness-associated mitochondrial tRNA(His) 12201T>C mutation. The destabilization of a highly conserved base-pairing (5A-68U) by the m.12201T>C mutation alters structure and function of tRNA(His). Using cybrids constructed by transferring mitochondria from lymphoblastoid cell lines derived from a Chinese family into mtDNA-less (rho(o)) cells, we showed similar to 70% decrease in the steady-state level of tRNA(His) in mutant cybrids, compared with control cybrids. The mutation changed the conformation of tRNA(His), as suggested by slower electrophoretic mobility of mutated tRNA with respect to the wild-type molecule. However, similar to 60% increase in aminoacylated level of tRNA(His) was observed in mutant cells. The failure in tRNA(His) metabolism was responsible for the variable reductions in seven mtDNA-encoded polypeptides in mutant cells, ranging from 37 to 81%, with the average of similar to 46% reduction, as compared with those of control cells. The impaired mitochondrial translation caused defects in respiratory capacity in mutant cells. Furthermore, marked decreases in the levels of mitochondrial ATP and membrane potential were observed in mutant cells. These mitochondrial dysfunctions caused an increase in the production of reactive oxygen species in the mutant cells. The data provide the evidence for a mitochondrial tRNA(His) mutation leading to deafness
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Background: Hypertrophic cardiomyopathy (HCM) is a primary disorder characterised by asymmetric thickening of septum and left ventricular wall, with a prevalence of 0.2% in the general population. Objective: To describe a novel mi...
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Background: Hypertrophic cardiomyopathy (HCM) is a primary disorder characterised by asymmetric thickening of septum and left ventricular wall, with a prevalence of 0.2% in the general population. Objective: To describe a novel mitochondrial DNA mutation and its association with the pathogenesis of HCM. Methods and results: All maternal members of a Chinese family with maternally transmitted HCM exhibited variable severity and age at onset, and were implanted permanent pacemakers due to complete atrioventricular block (AVB). Nuclear gene screening (MYH7, MYBPC3, TNNT2 and TNNI3) was performed, and no potential pathogenic mutation was identified. Mitochondrial DNA sequencing analysis identified a novel homoplasmic 16S rRNA 2336T>C mutation. This mutation was exclusively present in maternal members and absent in non-maternal members. Conservation index by comparison to 16 other vertebrates was 94.1%. This mutation disturbs the 2336U-A2438 base pair in the stem-loop structure of 16S rRNA domain III, which is involved in the assembly of mitochondrial ribosome. Oxygen consumption rate of the lymphoblastoid cells carrying 2336T>C mutation had decreased by 37% compared with controls. A reduction in mitochondrial ATP synthesis and an increase in reactive oxidative species production were also observed. Electron microscopic analysis indicated elongated mitochondria and abnormal mitochondrial cristae shape in mutant cells. Conclusions: It is suggested that the 2336T>C mutation is one of pathogenic mutations of HCM. This is the first report of mitochondrial 16S rRNA 2336T>C mutation and an association with maternally inherited HCM combined with AVB. Our findings provide a new insight into the pathogenesis of HCM.
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The brine shrimp, Anemia, reproduces either oviparously, producing encysted embryos (diapause cysts), or ovoviviparously, producing free-swimming nauplii. Environmental factors, such as photoperiod, have been applied to control th...
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The brine shrimp, Anemia, reproduces either oviparously, producing encysted embryos (diapause cysts), or ovoviviparously, producing free-swimming nauplii. Environmental factors, such as photoperiod, have been applied to control the reproduction mode of Artemia, but when the determination of a reproductive mode occurs remains unknown. We analysed the differential gene expression between oocytes from oviparous and ovoviviparous Artemia reared under different photoperiods. A total of 692 qualified cDNA clones were obtained by subtractive hybridization, 327 of which matched GenBank? Nucleotide Sequence Database entries. Gene expressions of 44 cDNAs (representing 56 clones) were analysed in oocytes using real-time PCR. Among these genes, 11 (21 clones) were significantly (P < 0.05) up-regulated and 7 (9 clones) down-regulated in Artemia oocytes that subsequently enter diapause. Remarkably, known diapause-related proteins such as ArHsp22 (Artemia heat-shock protein 22) and chitin-binding proteins are found to be already differentially expressed. Furthermore, RNAi (RNA interference) knockdown of a differentially expressed gene, polo-like kinase 1, in oocyte of ovoviviparous Artemia led to the production of white embryos rather than free-swimming nauplii. In summary, our results provide evidence at the molecular level that the reproductive mode of Artemia is already determined at the oocyte stage of their life cycle.
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The plant hormone auxin modulates cell proliferation and cell expansion in part by changing gene expression. Among the three primary auxin response gene families, Aux/IAA, GH3 and SAUR, the function of SAUR genes remains unclear. ...
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The plant hormone auxin modulates cell proliferation and cell expansion in part by changing gene expression. Among the three primary auxin response gene families, Aux/IAA, GH3 and SAUR, the function of SAUR genes remains unclear. SAUR transcripts were initially identified in epidermal and cortical cells of elongating tissues and thus were supposed to regulate cell expansion downstream of auxin transport and auxin signaling. Recent studies have proposed that SAUR proteins are able to modulate auxin transport and cell expansion by an unknown mechanism. We present our work on the SAUR41 subfamily genes of Arabidopsis (SAUR41, SAUR40, SAUR71 and SAUR72). Similar to the fusion protein between SAUR41 and EGFP, both SAUR40-EGFP and SAUR71-EGFP were identified in the cytoplasm of all types of root tip cells. This result indicated that the subcellular location pattern of SAUR proteins among the members of the same subfamily could be similar to each other, although the overall location pattern of SAUR proteins appeared to be highly diverse. SAUR41 was distinctively expressed in the quiescent center and cortex/endodermis initials of root stem cell niches and in the endodermis of hypocotyls, whereas SAUR71 and SAUR72 were expressed in the steles of young roots and hypocotyls. In addition, SAUR71 was differentially expressed during stomatal formation. The tissue-specific and developmentally regulated expression patterns of the SAUR41 subfamily genes imply that SAUR transcripts or SAUR proteins might serve as signal molecules to ensure the coordination of cell proliferation and cell expansion. Finally, Arabidopsis seedlings expressing TAP (tandem affinity peptide) tagged SAUR41 displayed phenotypes, indicating that it was rational to use the TAP approach for identification of potential binding partners of SAUR41 proteins.
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Hypertrophic cardiomyopathy is a primary disorder characterized by asymmetric thickening of the septum and left ventricular wall, which affects 1 in 500 individuals in the general population. Mutations in mitochondrial DNA have be...
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Hypertrophic cardiomyopathy is a primary disorder characterized by asymmetric thickening of the septum and left ventricular wall, which affects 1 in 500 individuals in the general population. Mutations in mitochondrial DNA have been found to be one of the most important causes of hypertrophic cardiomyopathy. Here we report the clinical, genetic and molecular characterization of a Han Chinese family with a likely maternally transmitted hypertrophic cardiomyopathy. Four (2 men/2 women) of 5 matrilineal relatives in this 3-generation family exhibited the variable severity and age at onset of 44 to 79. years old. Sequence analysis of the entire mitochondrial DNA in this pedigree identified the known homoplasmic . ND5 12338T>C variant. This mitochondrial DNA haplogroup belongs to the Eastern Asian F2a. The 12338T>C variant, highly evolutionarily conserved, resulted in the replacement of the translation initiating methionine with a threonine, shortening the . ND5 polypeptide by 2 amino acids. The occurrence of . ND5 12338T>C variant exclusively in maternal members of this Chinese family suggested that the 12338T>C variant is associated with maternally inherited hypertrophic cardiomyopathy. Our findings will provide theoretical basis for genetic counseling of maternally inherited hypertrophic cardiomyopathy.
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In crustaceans, the fascinating processes of maturation, reproductive molting and carapace coloration are regulated by hydrophobic molecules. Interestingly, most of the molecules are ligands of lipocalin. To understand the role of...
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In crustaceans, the fascinating processes of maturation, reproductive molting and carapace coloration are regulated by hydrophobic molecules. Interestingly, most of the molecules are ligands of lipocalin. To understand the role of lipocalin in the aforementioned processes at molecular level, we isolated a cDNA that belongs to the lipocalin family, from a central nervous system cDNA library of Macrobrachium rosenbergii. We monitored the spatial and temporal distributions of the mRNA by using Northern Blotting analysis. Our results demonstrated that this gene expresses abundantly in the subepidermal adipose tissue, while faintly in the hepatopancreas and central nervous system. However, no signal was detected in other tissues including muscle, gill and ovary. Its expression levels in subepidermal adipose tissue during various stages of maturation as well as through the whole molting cycle showed that prawn lipocalin is involved in sexual maturation, as the maximal level was observed just after molt.
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A protein encoded by At1g32080 was consistently identified in proteomic studies of Arabidopsis chloroplast envelope membranes, but its function remained unclear. The protein, designated AtLrgB, may have evolved from a gene fusion ...
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A protein encoded by At1g32080 was consistently identified in proteomic studies of Arabidopsis chloroplast envelope membranes, but its function remained unclear. The protein, designated AtLrgB, may have evolved from a gene fusion of IrgA and IrgB. Inbacteria, two homologous operons, IrgAB and cidAB, participate in an emerging mechanism to control cell death and lysis. We aim to characterize AtLrgB using reverse genetics and cell biological and biochemical analysis. AtLrgB is expressed in leaves, butnot in roots. T-DNA insertion mutation of AtLrgB produced plants with interveinal chlorotic and premature necrotic leaves. Overexpression of full-length AtLrgB (or its LrgA and LrgB domains, separately), under the control of CaMV 35S promoter, producedplants exhibiting veinal chlorosis and delayed greening. At the end of light period, the T-DNA mutant had high starch and low sucrose contents in leaves, while the 35S:AtLrgB plants had low starch and high sucrose contents. Metabolite profiling revealedthat AtLrgB appeared not to directly transport triose phosphate or hexose phosphates. In yeast cells, AtLrgB could augment nystatin-induced membrane permeability. Our work indicates that AtLrgB is a new player in chloroplast development, carbon partitioning and leaf senescence, although its molecular mechanism remains to be established.
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The thermo-responsivity of PNIPAAm [poly(N-isopropylcarylamide)]-grafted PDMS [poly(dimethylsiloxane)] surface is a property that could be feasibly used for detaching cells adhered on the surface. We used benzophenone-initiated ph...
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The thermo-responsivity of PNIPAAm [poly(N-isopropylcarylamide)]-grafted PDMS [poly(dimethylsiloxane)] surface is a property that could be feasibly used for detaching cells adhered on the surface. We used benzophenone-initiated photopolymerization to graft PNIPAAm on PDMS substrates to construct the PNIPAAm-grafted PDMS surface and this PDMS surface was highly thermo-responsive. hMSCs (human mesenchymal stem cells) were used to analyse the proliferation and multi-differentiation of stem cells on the PNIPAAm-grafted PDMS surface. The results showed that hMSCs could adhere on the PNIPAAm-grafted PDMS surface at 37 degrees C and form cell colonies, and then become fibroblastic. The proliferation potential of hMSCs on the PNIPAAm-grafted PDMS surface was not significantly different from that on a plate surface coated with gelatin. However, as it proved easier to detach cells from the surface, by changing temperature, a higher viability of detached cells could be obtained with the PNIPAAm-grafted PDMS surface, using a temperature shift, compared with a gelatin-coated surface, where cells are detached by treatment with trypsin. hMSCs on the PNIPAAm-grafted PDMS surface were induced into osteoblasts, adipocytes and neurocytes under osteogenic medium, adipogenic medium and neurogenic medium respectively. The PNIPAAm-grafted PDMS surface was favourable for osteogenesis of hMSCs, although the potentials of adipogenesis and neurogenesis of hMSCs on the PNIPAAm-grafted PDMS surface were similar to those on the plate surface coated with gelatin. The above results demonstrate that the PNIPAAm-grafted PDMS surface not only kept the potentials of proliferation and multi-differentiation of hMSCs, but also increased the viability of hMSCs.
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Under stress conditions transcription factors, including their coactivators, play major roles in mitochondrial biogenesis and oxidative phosphorylation. MED1 (Mediator complex subunit 1) functions as a coactivator of several trans...
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Under stress conditions transcription factors, including their coactivators, play major roles in mitochondrial biogenesis and oxidative phosphorylation. MED1 (Mediator complex subunit 1) functions as a coactivator of several transcription factors and is implicated in adipogenesis of the lipid and glucose metabolism. This suggests that MED1 may play a role in mitochondrial function. In this study, we found that both the mtDNA content and mitochondrial mass were markedly increased and cell proliferation markedly suppressed in MED1-deficient cells. Upon MED1 loss, Nrf1 and its downstream target genes involved in mitochondrial biogenesis (Tfam, Plormt, Tfb1m), were up-regulated as were those genes in the OXPHOS pathway. Moreover, the knockdown of MED1 resulted in significant changes in the profile of mitochondrial respiration, accompanied by a prominent decrease in the generation of ATP. Collectively, these observations strongly suggest that MED1 has an important affect on mitochondrial function. This further elucidates the role of MED1, particularly its role in the energy metabolism.
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It has been verified that placenta contains multi-lineage mesenchymal stem cells (MSCs). We have used a time-gradient attachment method to isolate placenta-derived MSCs (PMSCs). The morphology, differentiation potential, immunogen...
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It has been verified that placenta contains multi-lineage mesenchymal stem cells (MSCs). We have used a time-gradient attachment method to isolate placenta-derived MSCs (PMSCs). The morphology, differentiation potential, immunogenicity and xenogenic reconstruction potential of these PMSCs were examined. The results showed that PMSCs isolated using the time-gradient attachment method showed higher potential of in vitro proliferation and multi-lineage differentiation. PMSCs isolated using the time-gradient attachment method showed a low immunogenicity. HLA-A gene fragment and no HLA-DR gene fragment were detected in PMSCs isolated using the time-gradient attachment method, and the mixed lymphocyte reaction (MLR) assay identified that these cells inhibited the proliferation of the allogeneic T-lymphocytes induced by PHA. The transplantation in calvaria of rats showed that PMSCs had the higher xenogenic reconstruction potential. Finally, the significance of PMSCs isolated using the time-gradient attachment method in experimental and clinical applications is discussed.
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