The budding yeast Saccharomyces cerevisiae is an excellent model organism for studying chromatin regulation with high-resolution genome-wide analyses. Since newly generated genome-wide data are often compared with publicly available datasets, expanding our dataset repertoire will be beneficial for the field. Information on transcription start sites (TSSs) determined at base pair resolution is essential for elucidating mechanisms of transcription and related chromatin regulation, yet no datasets that cover two different cell types are available. Here, we present a CAGE (cap analysis of gene expression) dataset for a-cells and α-cells grown in defined and rich media. Cell type-specific genes were differentially expressed as expected, ensuring the reliability of the data. Some of the differentially expressed TSSs were medium-specific or detected due to unrecognized chromosome rearrangement. By comparing the CAGE data with a high-resolution nucleosome map, major TSSs were primarily found in +1 nucleosomes, with a peak approximately 30 bp from the promoter-proximal end of the nucleosome. The dataset is available at DDBJ/GEA.

The developmental mechanisms of limb buds have been studied in developmental biology as an excellent model of pattern formation. Chick embryos have contributed to the discovery of new principles in developmental biology, as it is easy to observe live embryos and manipulate embryonic tissues. Herein, I outline recent findings and future issues over the next decade regarding three themes, based on my research: limb positioning, proximal-distal limb elongation and digit identity determination. First, how hindlimb position is determined at the molecular level is described, with a focus on the transforming growth factor-β signaling molecule GDF11. Second, I explain how the cell population in the limb bud deforms with developmental progress, shaping the limb bud with elongation along the proximal-distal axis. Finally, I describe the developmental mechanisms that determine digit identity through the interdigits.

Response regulators (RRs) are implicated in various developmental processes as well as environmental responses by acting as either positive or negative regulators, and are crucial components of cytokinin signaling in plants. We characterized 36 RRs in rice (Oryza sativa L.; Os) using in silico analysis of publicly available data. A comprehensive analysis of OsRR family members covered their physicochemical properties, chromosomal distribution, subcellular localization, phylogeny, gene structure, distribution of conserved motifs and domains, and gene duplication events. Gene Ontology analysis indicated that 22 OsRR genes contribute mainly to the cytokinin response and signal transduction. Predicted cis-elements in RR promoter sequences related to phytohormones and abiotic stresses indicated that RRs are involved in hormonal and environmental responses, supporting previous studies. MicroRNA (miRNA) target analysis showed that 148 miRNAs target 29 OsRR genes. In some cases, multiple RRs are targets of the same miRNA group, and may be controlled by common stimulus responses. Based on the analysis of publicly available gene expression data, OsRR4, OsRR6, OsRR9, OsRR10, OsRR22, OsPRR73 and OsPRR95 were found to be involved in responses to abiotic stresses. Using quantitative reverse transcription polymerase chain reaction we confirmed that six of these RRs, namely OsRR4, OsRR6, OsRR9, OsRR10, OsRR22 and OsPRR73, are involved in the response to salinity, osmotic, alkaline and wounding stresses, and can potentially be used as models to understand molecular mechanisms underlying stress responsiveness.

We investigated the variation and geographical distribution of the Pseudo-regulator response 37 (Setaria italica PRR37; SiPRR37) gene, which is involved in heading time (photoperiodism) in foxtail millet. An allele of the SiPRR37 gene, in which an approximately 4.9-kb transposable element (designated TE1) is inserted (a loss-of-function or reduction-of-function type), is distributed sporadically in East Asia and broadly in Southeast Asia and South Asia, implying that this gene is important in latitudinal adaptation. In addition, we found a new allele of SiPRR37 with an insertion of a 360-bp TE (TE2) at this locus and investigated the geographical distribution of this new type. This SiPRR37 allele with TE2 is distributed in Japan, Korea, Nepal, Iran and Turkey. Both TE1 and TE2 are useful markers for tracing foxtail millet dispersal pathways in Asia.

Strict control of the expression levels of heterologously introduced protein-coding genes is important for the functional analysis of the protein of interest and its effective use in new situations. For this purpose, various promoters with different expression strengths, codon optimization, and expression stimulation by low-molecular-weight compounds are commonly used. However, methods to control protein expression levels by combining regulation of translation efficiency have not been studied in detail. We previously observed relatively high basal expression of Cre when it was heterologously expressed in fission yeast. Here, we used a fission yeast strain that is susceptible to centromere disruption, and thus highly sensitive to Cre levels, and report successful fine-tuning of heterologous Cre expression by modulating the Cre translation efficiency. To inhibit Cre translation initiation, we generated two mutations in the 5' untranslated region of the Cre mRNAs, both of which interfered with the scanning process of start codon recognition, mediated by specialized ribosomal subunits. These mutations successfully reduced the levels of exogenously expressed Cre to different degrees in fission yeast. Combining them with promoters of different strengths allowed us to conduct centromere disruption experiments in fission yeast. Our data indicate that modification of translational control is an additional tool in heterologous gene expression.

Homeostasis is essential for muscle repair and regeneration after skeletal muscle exercise. This study investigated the role of methyltransferase-like 21C (METTL21C) in skeletal muscle of mice after exercise and the potential mechanism. First, muscle samples were collected at 2, 4 and 6 weeks after exercise, and liver glycogen, muscle glycogen, blood lactic acid and triglyceride were assessed. Moreover, the expression levels of autophagy markers and METTL21C in skeletal muscle were analyzed. The results showed that the expression levels of METTL21C and MYH7 in the gastrocnemius muscle of mice in the exercise group were significantly higher after exercise than those in the control group, which suggested that long-term exercise promoted the formation of slow-twitch muscle fibers in mouse skeletal muscle. Likewise, the autophagy capacity was enhanced with the prolongation of exercise in muscles. The findings were confirmed in mouse C2C12 cells. We discovered that knockdown of Mettl21c reduced the expression of MYH7 and the autophagy level in mouse myoblasts. These findings indicate that METTL21C promotes skeletal muscle homeostasis after exercise by enhancing autophagy, and also contributes to myogenic differentiation and the formation of slow muscle fibers.

Lack of pigmentation in seed coats of soybean is caused by natural RNA silencing of chalcone synthase (CHS) genes. This phenomenon is an evolutionary consequence of structural changes in DNA that resulted in the production of double-stranded RNAs (dsRNAs) that trigger RNA degradation. Here we determined that a mutant with pigmented seed coats derived from a cultivar that lacked the pigmentation had a deletion between DNA regions ICHS1 and a cytochrome P450 gene; the deletion included GmIRCHS, a candidate gene that triggers CHS RNA silencing via production of CHS dsRNAs. We also characterized CHS short interfering RNAs (siRNAs) produced in the wild-type seed coats that had CHS RNA silencing. Phased 21-nt CHS siRNAs were detected in all 21 phases and were widely distributed in exon 2 of CHS7, which indicates commonality in the pattern of RNA degradation in natural CHS RNA silencing between distantly related species. These results with the similarities in the rearrangements found in spontaneous mutants suggest that the structural organization that generates dsRNAs that trigger phased siRNA production is vulnerable to further structural changes, which eventually abolish the induction of RNA silencing.

The importance of the parent-progeny relationship tracking technique in single-cell analysis has grown with the passage of time. In this study, fundamental image-processing techniques were combined to develop software capable of inferring cell cycle alterations in fission yeast cells, which exhibit equipartition during division. These methods, exclusively relying on bright-field images as input, could track parent-progeny relationships after cell division by assessing the temporal morphological transformation of these cells. In the application of this technique, the software was employed for calculating intracellular fluorescent dots during every stage of the cell cycle, using a yeast strain expressing EGFP-fused Swi6, which binds to chromatin. The results obtained with this software were consistent with those of previous studies. This software facilitated single-cell-level tracking of parent-progeny relationships in cells exhibiting equipartition during division and enabled the monitoring of spatial fluctuations in a cell cycle-dependent protein. This method, expediting the analysis of extensive datasets, may also empower large-scale screening experiments that cannot be conducted manually.

Mycoplasmas, autonomously culturable bacteria with the smallest genome, are an important organism to understand the minimal form of life. Mutagenesis using mutagens is a useful methodology for understanding the essential regions of genomic information. Ultraviolet light (UV) and trimethyl psoralen (TMP) are mutagens known to induce various mutations; the latter is reported to specifically induce deletions in nematodes. However, their mutagenic effects on mycoplasma are not known. Here, we exposed Metamycoplasma salivarium to UV-C light or TMP and UV-A as mutagens, and analyzed the mutational pattern after serial cultivation ranging from 34 to 56 rounds for different lineages. Our results showed that more deletions, but fewer point mutations, were induced with TMP and UV-A than with UV-C, indicating the usefulness of TMP in inducing deletions. In addition, we compared our results with mutational data from other studies, which suggested that the combination of TMP and UV-A or UV-C exposure both induced point mutations that were highly biased toward C→T and G→A transitions. These data provide useful basic knowledge for mutational studies on M. salivarium.

Next-generation sequencing (NGS) has become widely available and is routinely used in basic research and clinical practice. The reference genome sequence is an essential resource for NGS analysis, and several population-specific reference genomes have recently been constructed to provide a choice to deal with the vast genetic diversity of human samples. However, resources supporting population-specific references are insufficient, and it is burdensome to perform analysis using these reference genomes. Here, we constructed a set of resources to support NGS analysis using the Japanese reference genome, JG. We created resources for variant calling, variant effect prediction, gene and repeat element annotations, read mappability and RNA-seq analysis. We also provide a resource for reference coordinate conversion for further annotation enrichment. We then provide a variant calling protocol with JG. Our resources provide a guide to prepare sufficient resources for the use of population-specific reference genomes and can facilitate the migration of reference genomes.