Genes to Cells最新文献

We previously developed a genome engineering method (TAQing2.0) based on the direct delivery of DNA endonucleases into living cells, which induces genome rearrangements even in non-sporulating nonconventional yeasts without introducing foreign DNA. Using TAQing2.0 and conventional mutagenesis (by nitrosoguanidine), we obtained mutant asexual Candida utilis strains capable of growing under highly acidic conditions (pH 1.8). Whole genome resequencing revealed that the genomic sequences of mutants generated by both methods contain a negligible small population of unmappable sequences, suggesting that both types of mutants can be regarded as equivalent to naturally occurring mutants. TAQing2.0 mutants exhibit multiple genome rearrangements with few point mutations, whereas conventional mutagenesis produces numerous point mutations. This feature enabled us to easily identify candidate genes (e.g., LYP1 homolog) responsible for acid resistance. TAQing2.0 is a powerful and versatile tool for mutant production and gene hunting without invasion of foreign DNA.

Naito, A., S. Kamakura, J. Hayase, et al. 2025. “The Protein Kinase aPKC as Well as the Small GTPases RhoA and Cdc42 Regulates Neutrophil Chemotaxis Partly by Recruiting the ROCK Kinase to the Leading Edge.” Genes to Cells 30, no. 2: e70002. https://doi.org/10.1111/gtc.70002.

In the “Funding” section on page 1, the text “This work was supported by Japan Society for the Promotion of Science (JP21H02698, JP21H05267, and JP22K06901).” was incorrect. This should have read: “This work was supported by Japan Society for the Promotion of Science (JP21H02698, JP21H05267, and JP22K06901) and by the fellowship from the Takeda Science Foundation.”

In the “Acknowledgments” section on page 17, the text “This work was supported in part by JSPS (Japan Society for the Promotion of Science) KAKENHI Grants JP21H02698 (to H.S.), JP21H05267 (to H.S.), and JP22K06901 (to S.K.).” was incorrect. This should have read: “This work was supported in part by JSPS (Japan Society for the Promotion of Science) KAKENHI Grants JP21H02698 (to H.S.), JP21H05267 (to H.S.), and JP22K06901 (to S.K.), and by the fellowship from the Takeda Science Foundation (to A.N.).”

We apologize for these errors.

Human sapovirus (HuSaV), first identified in the 1970s, is a significant cause of acute gastroenteritis, particularly in young children. Despite its clinical significance, research on HuSaV has been limited due to the absence of a reliable cell culture system. In 2020, a breakthrough study reported that HuSaV GI.1 and GII.3 strains could be cultured and serially propagated using HuTu80 cells in the presence of bile acids. However, in 2024, a subsequent study reported that effective replication in HuTu80 cells requires specialized cells that have undergone over 100 passages. In this study, we sought to identify an alternative cell culture system for HuSaV. HuSaV GI.1 can replicate and be serially propagated using Caco-2 cells under bile acid supplementation. Importantly, the Caco-2 cells were freshly sourced from the American Type Culture Collection, ensuring reproducibility for laboratories worldwide. Furthermore, Caco-2MC cells were established via single-cell cloning from in-house Caco-2/Cas9 cells with 91.5% HuSaV-susceptible. HuSaV strains GI.1, GI.2, GI.3, GII.1, GII.3, and GV.1 were successfully propagated using Caco-2MC cells, with RNA copy numbers increasing up to 4.4 log₁₀-fold within 5 days post-infection. This efficient HuSaV cell culture system represents a significant advancement in HuSaV research.

Retinal ischemic disease results in significant visual impairment due to the development of fragile and disorganized, pathologically running blood vessels in the eye. Currently, the mainstay treatment for this disease is the intravitreal administration of anti-VEGF drugs targeting vascular endothelial growth factor (VEGF), which induces angiogenesis. However, current anti-VEGF drugs do not diminish the ischemic areas that lead to angiogenesis, making fundamental treatment challenging. Since retinopathy is an acquired disease caused by hypoxic stimulation from ischemia, we paid particular attention to histone acetylases. We conducted a drug screening experiment using a mouse model of oxygen-induced retinopathy (OIR), which replicates retinal ischemic disease, through the intraperitoneal administration of 17 distinct inhibitors targeting histone acetyltransferases (HAT). The results indicated that, among the 17 inhibitors, only ISOX-DUAL decreased neovascularization and ischemic regions. Furthermore, microarray analysis was conducted on the drug-treated samples to refine genes altered by the administration of ISOX-DUAL. There were 21 genes associated with angiogenesis, including Angpt2, Hmox1, Edn1, and Serpine1, exhibited upregulation in OIR mice and downregulation following treatment with ISOX-DUAL. Furthermore, STRING analysis confirmed that the aforementioned four genes are downstream factors of hypoxia-inducible factors and are assumed to be important factors in retinal ischemic diseases.

The small GTPases RhoA and Cdc42 and their effector proteins play crucial roles in neutrophil chemotaxis. However, endogenous localization and regulation of these proteins have remained largely unknown. Here, we show, using a trichloroacetic acid fixation method, that endogenous RhoA and Cdc42 are preferentially accumulated at the F-actin-rich leading edge (pseudopod) during chemotaxis of human neutrophil-like PLB-985 cells in response to the chemoattractant C5a. Interestingly, the enrichment of RhoA is impaired by knockdown of Cdc42, indicating a positive regulation by Cdc42. Depletion of Cdc42 or RhoA each induces the formation of multiple pseudopods, confirming their significance in cell polarization with an organized actin network at the front. The Rho-associated kinase ROCK is also recruited to the leading edge during chemotaxis in a manner dependent on not only RhoA and Cdc42 but also aPKC, a Cdc42-interacting kinase that can also bind to ROCK. ROCK promotes phosphorylation of the myosin light chain at the front, possibly regulating pseudopod contractility. Knockdown of aPKC suppresses neutrophil chemotaxis by disturbing pseudopod orientation without forming multiple protrusions. An incorrectly oriented pseudopod is also observed in ROCK-depleted cells. Thus, aPKC, as well as RhoA and Cdc42, likely regulates neutrophil chemotaxis partly by recruiting ROCK to the leading edge for correct directionality.