In the past decade, the development of next-generation sequencing (NGS) has paved the way for whole-genome analysis in individuals. Research on the human leukocyte antigen (HLA), an extensively ...studied molecule involved in immunity, has benefitted from NGS technologies. The HLA region, a 3.6-Mb segment of the human genome at 6p21, has been associated with more than 100 different diseases, primarily autoimmune diseases. Recently, the HLA region has received much attention because severe adverse effects of various drugs are associated with particular HLA alleles. Owing to the complex nature of the HLA genes, classical direct sequencing methods cannot comprehensively elucidate the genomic makeup of HLA genes. Thus far, several high-throughput HLA-typing methods using NGS have been developed. In HLA research, NGS facilitates complete HLA sequencing and is expected to improve our understanding of the mechanisms through which HLA genes are modulated, including transcription, regulation of gene expression and epigenetics. Most importantly, NGS may also permit the analysis of HLA-omics. In this review, we summarize the impact of NGS on HLA research, with a focus on the potential for clinical applications.
The human leukocyte antigen (HLA) super-locus is a genomic region in the chromosomal position 6p21 that encodes the six classical transplantation HLA genes and at least 132 protein coding genes that ...have important roles in the regulation of the immune system as well as some other fundamental molecular and cellular processes. This small segment of the human genome has been associated with more than 100 different diseases, including common diseases, such as diabetes, rheumatoid arthritis, psoriasis, asthma and various other autoimmune disorders. The first complete and continuous HLA 3.6 Mb genomic sequence was reported in 1999 with the annotation of 224 gene loci, including coding and non-coding genes that were reviewed extensively in 2004. In this review, we present (1) an updated list of all the HLA gene symbols, gene names, expression status, Online Mendelian Inheritance in Man (OMIM) numbers, including new genes, and latest changes to gene names and symbols, (2) a regional analysis of the extended class I, class I, class III, class II and extended class II subregions, (3) a summary of the interspersed repeats (retrotransposons and transposons), (4) examples of the sequence diversity between different HLA haplotypes, (5) intra- and extra-HLA gene interactions and (6) some of the HLA gene expression profiles and HLA genes associated with autoimmune and infectious diseases. Overall, the degrees and types of HLA super-locus coordinated gene expression profiles and gene variations have yet to be fully elucidated, integrated and defined for the processes involved with normal cellular and tissue physiology, inflammatory and immune responses, and autoimmune and infectious diseases.
Induced pluripotent stem cells (iPSCs) constitute a potential source of autologous patient-specific cardiomyocytes for cardiac repair, providing a major benefit over other sources of cells in terms ...of immune rejection. However, autologous transplantation has substantial challenges related to manufacturing and regulation. Although major histocompatibility complex (MHC)-matched allogeneic transplantation is a promising alternative strategy, few immunological studies have been carried out with iPSCs. Here we describe an allogeneic transplantation model established using the cynomolgus monkey (Macaca fascicularis), the MHC structure of which is identical to that of humans. Fibroblast-derived iPSCs were generated from a MHC haplotype (HT4) homozygous animal and subsequently differentiated into cardiomyocytes (iPSC-CMs). Five HT4 heterozygous monkeys were subjected to myocardial infarction followed by direct intra-myocardial injection of iPSC-CMs. The grafted cardiomyocytes survived for 12 weeks with no evidence of immune rejection in monkeys treated with clinically relevant doses of methylprednisolone and tacrolimus, and showed electrical coupling with host cardiomyocytes as assessed by use of the fluorescent calcium indicator G-CaMP7.09. Additionally, transplantation of the iPSC-CMs improved cardiac contractile function at 4 and 12 weeks after transplantation; however, the incidence of ventricular tachycardia was transiently, but significantly, increased when compared to vehicle-treated controls. Collectively, our data demonstrate that allogeneic iPSC-CM transplantation is sufficient to regenerate the infarcted non-human primate heart; however, further research to control post-transplant arrhythmias is necessary.
Recent studies have demonstrated that chloroplasts and mitochondria evoke specific Ca2＋ signals in response to biotic and abiotic stresses in a stress-dependent manner. The identification of Ca2＋ ...transporters and Ca2＋signaling mol- ecules in chloroplasts and mitochondria implies that they play roles in controlling not only intra-organellar functions, but also extra-organellar processes such as plant immunity and stress responses. It appears that organellar Ca2＋ signaling might be more important to plant cell functions than previously thought. This review briefly summarizes what is known about the molecular basis of Ca2＋ signaling in plant mitochondria and chloroplasts.
Chloroplasts import thousands of nucleus-encoded preproteins synthesized in the cytosol through the TOC and TIC translocons on the outer and inner envelope membranes, respectively. Preprotein ...translocation across the inner membrane requires ATP; however, the import motor has remained unclear. Here, we report that a 2-MD heteromeric AAA-ATPase complex associates with the TIC complex and functions as the import motor, directly interacting with various translocating preproteins. This 2-MD complex consists of a protein encoded by the previously enigmatic chloroplast gene
and five related nuclear-encoded FtsH-like proteins, namely, FtsHi1, FtsHi2, FtsHi4, FtsHi5, and FtsH12. These components are each essential for plant viability and retain the AAA-type ATPase domain, but only FtsH12 contains the zinc binding active site generally conserved among FtsH-type metalloproteases. Furthermore, even the FtsH12 zinc binding site is dispensable for its essential function. Phylogenetic analyses suggest that all AAA-type members of the Ycf2/FtsHi complex including Ycf2 evolved from the chloroplast-encoded membrane-bound AAA-protease FtsH of the ancestral endosymbiont. The Ycf2/FtsHi complex also contains an NAD-malate dehydrogenase, a proposed key enzyme for ATP production in chloroplasts in darkness or in nonphotosynthetic plastids. These findings advance our understanding of this ATP-driven protein translocation system that is unique to the green lineage of photosynthetic eukaryotes.
Chloroplasts are semiautonomous organelles which possess their own genome and gene expression system. However, extant chloroplasts contain only limited coding information, and are dependent on a ...large number of nucleus-encoded proteins. During plant evolution, chloroplasts have lost most of the prokaryotic DNA-binding proteins and transcription regulators that were present in the original endosymbiont. Thus, chloroplasts have a unique hybrid transcription system composed of the remaining prokaryotic components, such as a prokaryotic RNA polymerase as well as nucleus-encoded eukaryotic components. Recent proteomic and transcriptomic analyses have provided insights into chloroplast transcription systems and their evolution. Here, we review chloroplast-specific transcription systems, focusing on the multiple RNA polymerases, eukaryotic transcription regulators in chloroplasts, chloroplast promoters, and the dynamics of chloroplast nucleoids.
In bacteria, the hyper-phosphorylated nucleotide, guanosine 3',5'-bis(pyrophosphate) (ppGpp), functions as a secondary messenger under stringent conditions. ppGpp levels are controlled by two ...distinct enzymes, namely RelA and SpoT, in Escherichia coli. RelA-SpoT homologs (RSHs) are also conserved in plants where they function in the plastids. The model plant Arabidopsis thaliana contains four RSHs: RSH1, RSH2, RSH3 and Ca2+-dependent RSH (CRSH). Genetic characterizations of RSH1, RSH2 and RSH3 were undertaken, which showed that the ppGpp-dependent plastidial stringent response significantly influences plant growth and stress acclimation. However, the physiological significance of CRSH-dependent ppGpp synthesis remains unclear, as no crsh-null mutant has been available. Here, to investigate the function of CRSH, a crsh-knockout mutant of Arabidopsis was constructed using a site-specific gene-editing technique, and its phenotype was characterized. A transient increase in ppGpp was observed for 30 min in the wild type (WT) after the light-to-dark transition, but this increase was not observed in the crsh mutant. Similar analyses were performed with the rsh2-rsh3 double and rsh1-rsh2-rsh3 triple mutants and showed that the transient increments of ppGpp in the mutants were higher than those in the WT. The increase in ppGpp in the WT and rsh2 rsh3 accompanied decrements in the mRNA levels of some plastidial genes transcribed by the plastid-encoded plastid RNA polymerase. These results indicate that the transient increase in ppGpp at night is due to CRSH-dependent ppGpp synthesis and that the ppGpp level is maintained by the hydrolytic activities of RSH1, RSH2 and RSH3 to accustom plastidial gene expression to darkness.
Summary
The MHC is a highly polymorphic genomic region that encodes the transplantation and immune regulatory molecules. It receives special attention for genetic investigation because of its ...important role in the regulation of innate and adaptive immune responses and its strong association with numerous infectious and/or autoimmune diseases. The MHC locus was first discovered in the mouse and for the past 50 years it has been studied most intensively in both mice and humans. However, in recent years the macaque species have emerged as some of the more important and advanced experimental animal models for biomedical research into MHC with important human immunodeficiency virus/simian immunodeficiency virus and transplantation studies undertaken in association with precise MHC genotyping and haplotyping methods using Sanger sequencing and next‐generation sequencing. Here, in this special issue on ‘Macaque Immunology’ we provide a short review of the genomic similarities and differences among the human, macaque and mouse MHC class I and class II regions, with an emphasis on the association of the macaque class I region with MHC polymorphism, haplotype structure and function.
We have provided a short review of the genomic similarities and differences between the macaque, human and mouse MHC regions with a particular emphasis on the comparative polymorphisms, haplotype structure and function of the class I region of humans and macaques. The macaque species have had many more MHC class I genes generated by gene duplication events than those in humans, whereas the organization of MHC class II genes are well conserved between the two species.
Among the non-human primates used in experimental medicine, cynomolgus macaques (
hereafter referred to as
) are increasingly selected for the ease with which they are maintained and bred in ...captivity. Macaques belong to Old World monkeys and are phylogenetically much closer to humans than rodents, which are still the most frequently used animal model. Our understanding of the
genome has progressed rapidly in recent years and has greatly benefited from the latest technical advances in molecular genetics. Cynomolgus macaques are widespread in Southeast Asia and numerous studies have shown a distinct genetic differentiation of continental and island populations. The major histocompatibility complex of cynomolgus macaque (
MHC) is organized in the same way as that of human, but it differs from the latter by its high degree of classical class I gene duplication. Human polymorphic MHC regions play a pivotal role in allograft transplantation and have been associated with more than 100 diseases and/or phenotypes. The
MHC polymorphism similarly plays a crucial role in experimental allografts of organs and stem cells. Experimental results show that the
MHC class I and II regions influence the ability to mount an immune response against infectious pathogens and vaccines. MHC also affects cynomolgus macaque reproduction and impacts on numerous biological parameters. This review describes the
MHC polymorphism and the methods currently used to characterize it. We discuss some of the major areas of experimental medicine where an effect induced by MHC polymorphism has been demonstrated.
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