抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

Aminoacyl-tRNAs are substrates for translation and are pivotal in determining how the genetic code is interpreted as amino acids. The function of aminoacyl-tRNA synthesis is to precisely match amino acids with tRNAs containing the corresponding anticodon. This is primarily achieved by the direct attachment of an amino acid to the corresponding tRNA by an aminoacyl-tRNA synthetase, although intrinsic proofreading and extrinsic editing are also essential in several cases. Recent studies of aminoacyl-tRNA synthesis, mainly prompted by the advent of whole genome sequencing and the availability of a vast body of structural data, have led to an expanded and more detailed picture of how aminoacyl-tRNAs are synthesized. This article reviews current knowledge of the biochemical, structural, and evolutionary facets of aminoacyl-tRNA synthesis.

Aminoacyl-tRNA synthesis.

CRISPR-Cas9 is a versatile genome editing technology for studying the functions of genetic elements. To broadly enable the application of Cas9 in vivo, we established a Cre-dependent Cas9 knockin mouse. We demonstrated in vivo as well as ex vivo genome editing using adeno-associated virus (AAV)-, lentivirus-, or particle-mediated delivery of guide RNA in neurons, immune cells, and endothelial cells. Using these mice, we simultaneously modeled the dynamics of KRAS, p53, and LKB1, the top three significantly mutated genes in lung adenocarcinoma. Delivery of a single AAV vector in the lung generated loss-of-function mutations in p53 and Lkb1, as well as homology-directed repair-mediated KrasG12D mutations, leading to macroscopic tumors of adenocarcinoma pathology. Together, these results suggest that Cas9 mice empower a wide range of biological and disease modeling applications.

CRISPR-Cas9 Knockin Mice for Genome Editing and Cancer Modeling

A genetic annealing model for the universal ancestor of all extant life is presented; the name of the model derives from its resemblance to physical annealing. The scenario pictured starts when "genetic temperatures" were very high, cellular entities (progenotes) were very simple, and information processing systems were inaccurate. Initially, both mutation rate and lateral gene transfer levels were elevated. The latter was pandemic and pervasive to the extent that it, not vertical inheritance, defined the evolutionary dynamic. As increasingly complex and precise biological structures and processes evolved, both the mutation rate and the scope and level of lateral gene transfer, i.e., evolutionary temperature, dropped, and the evolutionary dynamic gradually became that characteristic of modern cells. The various subsystems of the cell "crystallized," i.e., became refractory to lateral gene transfer, at different stages of "cooling," with the translation apparatus probably crystallizing first. Organismal lineages, and so organisms as we know them, did not exist at these early stages. The universal phylogenetic tree, therefore, is not an organismal tree at its base but gradually becomes one as its peripheral branchings emerge. The universal ancestor is not a discrete entity. It is, rather, a diverse community of cells that survives and evolves as a biological unit. This communal ancestor has a physical history but not a genealogical one. Over time, this ancestor refined into a smaller number of increasingly complex cell types with the ancestors of the three primary groupings of organisms arising as a result.

https://www.pnas.org/content/pnas/95/12/6854.full.pdf

The universal ancestor

Correct expression of the genetic code at translation is directly correlated with tRNA identity. This survey describes the molecular signals in tRNAs that trigger specific aminoacylations. For most tRNAs, determinants are located at the two distal extremities: the anticodon loop and the amino acid accepting stem. In a few tRNAs, however, major identity signals are found in the core of the molecule. Identity elements have different strengths, often depend more on k cat effects than on K m effects and exhibit additive, cooperative or anti-cooperative interplay. Most determinants are in direct contact with cognate synthetases, and chemical groups on bases or ribose moieties that make functional interactions have been identified in several systems. Major determinants are conserved in evolution; however, the mechanisms by which they are expressed are species dependent. Recent studies show that alternate identity sets can be recognized by a single synthetase, and emphasize the importance of tRNA architecture and anti-determinants preventing false recognition. Identity rules apply to tRNA-like molecules and to minimalist tRNAs. Knowledge of these rules allows the manipulation of identity elements and engineering of tRNAs with switched, altered or multiple specificities.

/pdf/universal-rules-and-idiosyncratic-features-in-trna-identity-3onunm1vku.pdf

Universal rules and idiosyncratic features in tRNA identity

https://www.uab.edu/medicine/rheumatology/images/AG_-_Funabiki_et_al_2014.pdf

Autoimmune disorders associated with gain of function of the intracellular sensor MDA5.

Cytoplasmic aminoacyl-tRNA synthetases of higher eukaryotes acquired extra peptides in the course of their evolution. It has been thought that these appendices are related to the occurrence of the multiprotein complex consisting of at least eight different tRNA synthetase polypeptides. This complex is believed to be a signature feature of metazoans. In this study, we used multiple sequence alignments to infer the locations of the peptide appendices from human cytoplasmic tRNA synthetases found in the multisynthetase complex. The selected peptide appendices ranged from 22 aa of aspartyl-tRNA synthetase to 267 aa of methionyl-tRNA synthetase. We then made genetic constructions to investigate interactions between all 64 combinations of these peptides that were individually fused to nonsynthetase test proteins. The analyses identified 11 (10 heterologous and 1 homologous) interactions. The six peptide-dependent interactions paralleled what had been detected by crosslinking methods applied to the isolated multisynthetase complex. Thus, small peptide appendices seem to link together different synthetases into a complex. In addition, five interacting pairs that had not been detected previously were suggested from the observed peptide-dependent complexes.

https://www.pnas.org/content/pnas/96/8/4488.full.pdf

Genetic dissection of protein-protein interactions in multi-trna synthetase complex

Human glycyl-tRNA synthetase. Wide divergence of primary structure from bacterial counterpart and species-specific aminoacylation.

Cytoplasmic aminoacyl-tRNA synthetases of higher eukaryotes acquired extra peptides in the course of their evolution. It has been thought that these appendices are related to the occurrence of the multiprotein complex con- sisting of at least eight different tRNA synthetase polypep- tides. This complex is believed to be a signature feature of metazoans. In this study, we used multiple sequence align- ments to infer the locations of the peptide appendices from human cytoplasmic tRNA synthetases found in the multisyn- thetase complex. The selected peptide appendices ranged from 22 aa of aspartyl-tRNA synthetase to 267 aa of methionyl- tRNA synthetase. We then made genetic constructions to investigate interactions between all 64 combinations of these peptides that were individually fused to nonsynthetase test proteins. The analyses identified 11 (10 heterologous and 1 homologous) interactions. The six peptide-dependent interac- tions paralleled what had been detected by crosslinking methods applied to the isolated multisynthetase complex. Thus, small peptide appendices seem to link together different synthetases into a complex. In addition, five interacting pairs that had not been detected previously were suggested from the observed peptide-dependent complexes. Proteins are molecular fossils that may help to unravel the history and mechanism of evolution. Aminoacyl-tRNA syn- thetases (ARSs) are one of the ancient proteins evolved to decode genetic information to amino acids. Although all of the tRNA synthetases catalyze the same chemical reactions, these enzymes have accumulated a wide range of sequence and structural diversity throughout evolution (1). In particular, the tRNA synthetases of higher eukaryotes have acquired a few features that are not present in those of other organisms. Most intriguing is the presence of noncatalytic peptide appendices, the roles of which are yet to be elucidated. It has been thought that these peptides are involved in protein-protein interactions among the tRNA synthetases that form the multiprotein complex. This complex is another characteristic of the cyto- plasmic tRNA synthetases of higher eukaryotes (2-4) and contains at least eight different tRNA synthetase polypeptides (Glu-Pro-tRNA synthetase (EPRS), IRS, LRS, MRS, QRS, RRS, KRS, and DRS; refs. 5-8), as well as three nonsynthetase proteins of 43 kDa, 38 kDa, and 18 kDa (5-11). Although these complexes have been known for more than two decades, the structural organization and interactions between the components have not been well understood. Structural analysis with electron microscopy has shown that the complex forms an elongated U shape (8). The complex structure has been studied extensively by stepwise dissociation of the components by using nonionic detergent (12, 13), changing salt concentration (14), and chemical crosslinking (15, 16). The multi-ARS complex has been proposed to consist of three subdomains. The base subdomain consists of EPRS, IRS, and LRS, which are the enzymes of higher molecular weights. There are two arm subdomains on the top of the base subdomain. Subdomain I contains a dimer of DRS and mono- mers of MRS and QRS; subdomain II is made of dimers of KRS and RRS (16). Most of the structural analyses on the multi-ARS complex have relied on biochemical approaches, as listed above. Be- cause these methods address only the physical relationships of the protein components, other approaches are necessary to determine the molecular mechanisms and the peptide regions involved in the assembly or maintenance of the complex. We thought that a genetic approach would be suitable to investi- gate these questions. A yeast two-hybrid system has proven useful to analyze protein-protein interactions (17). This method was employed previously to analyze the interactions of the repeated motifs of EPRS with the C-terminal repeated motifs of IRS (18), as well as with the N-terminal extension of RRS (19). The interaction between the repeats of EPRS and IRS was confirmed further by biochemical and biophysical methods (19). Interactions of p38 with the other ARS com- ponents recently have been reported by using two-hybrid analysis (11). The role of the peptide appendices in the formation of the multi-ARS complex has been studied in a few cases such as the N-terminal sequences of DRS (20, 21), RRS (22, 23), and KRS (6). However, systematic studies on the structural features and molecular interactions of these peptides in eukaryotic ARSs are needed to understand their functional significance. In the present study, we focused on the peptide appendices deduced from the complex-forming ARSs. We investigated whether these regions actually are involved in protein-protein interac- tions among the complex-forming ARSs and, if so, how they are connected to each other. Finally, the interactions between the unique peptides of complex-forming tRNA synthetases determined by the genetic method in this work were compared with those previously obtained by biochemical methods.

Hiromi Motegi

Papers

Autoimmune disorders associated with gain of function of the intracellular sensor MDA5.

Genetic dissection of protein-protein interactions in multi-trna synthetase complex

Human glycyl-tRNA synthetase. Wide divergence of primary structure from bacterial counterpart and species-specific aminoacylation.

Genetic dissection of protein-protein interactions in multi-tRNA synthetase complex (peptide appendicesyyeast two-hybrid assayyinteraction map)

Precursor of pro-apoptotic cytokine modulates aminoacylation activity of tRNA synthetase.