Biology subjects

Kaku, M.

Publications and source records attributed to Kaku, M..

4 recordsLinked to original sources

bioRxiv2025

Ex vivo HIV DNA integration in STAT3 drives T cell persistence--A model of HIV-associated T cell lymphoma

Oncogenic retroviruses are known for their pathogenesis via insertional mutagenesis, in which the presence of a provirus and its transcriptional control elements alter the expression of a nearby or surrounding host gene. There are reports of proviral integration driving oncogenesis in people with HIV and the use of HIV-derived vectors for gene therapy has raised concern about oncogenic side effects. To study this issue, we used an ex vivo human CD4+ T cell infection model developed in our laboratory to identify HIV-1 integration sites that might influence cell proliferation or survival. Combining integration site analysis and bulk RNA sequencing, we established that an upregulated STAT3 signature due to proviral insertional mutagenesis was associated with persistent HIV-infected CD4+ T cells. HIV+ persistent cells also expressed a STAT3-related anti-apoptotic and cytotoxic phenotype that resembles that of HIV-associated T cell lymphomas. HIV insertional mutagenesis of STAT3 and expression of its downstream targets provides a model of HIV-associated T cell lymphomas that can be used to further determine the oncogenic drivers of HIV-associated lymphomas, both AIDS- and gene therapy-associated, and, potentially, to evaluate therapeutics against these HIV-associated cancers. Author SummaryThe effects of HIV proviral insertional mutagenesis have been demonstrated in a handful of HIV-associated T cell lymphomas, where integration of an HIV provirus within intron 1 of STAT3, results in increased expression of the STAT3 protein. To study the effects of HIV insertional mutagenesis, we established an ex vivo culture protocol of primary human CD4+ T cells infected with a replication-incompetent HIV vector with a gfp-reporter. After infection, the HIV/GFP+ cells from all three donors declined, but, over time, 3/6 replicates from one donor populations of infected cells rebounded. The resurgent HIV/GFP+ cells contained a provirus integrated within intron 1 of STAT3, which led to increases in gene expression, STAT3 activation, and upregulation of a STAT3-associated anti-apoptotic and cytotoxic phenotype. The STAT3-associated gene signature shared similarities to the HIV-associated lymphomas with similar integration sites. Additionally, in all 3 replicates, insertional mutagenesis of genes other than STAT3 may have also contributed to clonal expansion of HIV/GFP+ T cells. Overall, we have demonstrated that HIV provirus insertional mutagenesis can influence T cell persistence. Our study provides a primary T cell culture model system that can be used to further study how proviral insertional mutagenesis influences HIV-associated T cell lymphomas and the safety of lentiviral vectors used in gene and cell therapies.

microbiology↗

bioRxiv2024

Interferon-β induction heterogeneity during KSHV infection is determined by interferon-β enhanceosome transcription factors other than IRF3

Careful regulation of type I interferons (IFN) is vital for balancing tissue damage and protection against infections. We previously found that during Kaposis sarcoma-associated herpesvirus infection, type I IFN induction was limited to a small percentage of infected cells. This heterogeneity was not explained by viral gene expression. Here, we used a fluorescent reporter and fluorescence activated cell sorting to investigate the source of this heterogeneity. Surprisingly, the canonical IFN induction pathway culminating in the activation of the IRF3 transcription factor was similarly activated between cells that made high vs. low/no IFN-{beta}. In contrast, the activation or expression of the two other IFN transcription factors, the NF-{kappa}B subunit RelA and the AP-1 subunit ATF2, correlated with IFN-{beta} induction. Our results suggest that during viral infection, activation of IRF3 does not automatically result in IFN responses at the level of individual cells, but that other factors, such as NF-{kappa}B and AP-1, are limiting for type I IFN induction. ImportanceThe ability of mammalian cells to react to viral infections is a crucial step in the induction of immune responses. The first course of action for the cell is to express and release type I interferons like interferon-{beta} (IFN-{beta}), secreted molecules that warn surrounding cells. Single-cell level examination of gene expression has revealed that surprisingly, during many viral infections, only a small fraction of infected cells makes IFN-{beta}. This is likely a mechanism to prevent immune system overreactions. However, it remains unclear why only some cells respond. Here, we find that during infection with Kaposis sarcoma-associated herpesvirus, an oncogenic virus that affects immunocompromised individuals, the transcription factors AP-1 and NF-{kappa}B, rather than the more commonly studied IRF3, may decide which cells go on to make IFN-{beta}. Our findings contribute to a better understanding of complex gene regulation and shed light on a process that fights an oncogenic virus.

microbiology↗

bioRxiv2019

Development and validation of a deep learning framework for Alzheimers disease classification

Alzheimers disease (AD) is the primary cause of dementia worldwide (1), with an increasing morbidity burden that may outstrip diagnosis and management capacity as the population ages. Current methods integrate patient history, neuropsychological testing and magnetic resonance imaging (MRI) to identify likely cases, yet effective practices remain variably-applied and lacking in sensitivity and specificity (2). Here we report an explainable deep learning strategy that delineates unique AD signatures from multimodal inputs of MRI, age, gender, and mini-mental state examination (MMSE) score. Our framework linked a fully convolutional network (FCN) to a multilayer perceptron (MLP) to construct high resolution maps of disease probability from local brain structure. This enabled precise, intuitive visualization of individual AD risk en route to accurate diagnosis. The model was trained using clinically-diagnosed AD and cognitively normal (NC) subjects from the Alzheimers Disease Neuroimaging Initiative (ADNI) dataset (n=417) (3), and validated on three independent cohorts: the Australian Imaging, Biomarker & Lifestyle Flagship Study of Ageing (AIBL, n=382) (4), the Framingham Heart Study (FHS, n=102) (5), and the National Alzheimers Coordinating Center (NACC, n=582) (6). Model performance was consistent across datasets, with mean accuracy values of 0.966, 0.948, 0.815, and 0.916 for ADNI, AIBL, FHS and NACC, respectively. Moreover, our approach exceeded the diagnostic performance of a multi-institutional team of practicing neurologists (n=11), and high-risk cerebral regions predicted by the model closely tracked postmortem histopathological findings. This framework provides a clinically-adaptable strategy for using routinely available imaging techniques such as MRI to generate nuanced neuroimaging signatures for AD diagnosis, as well as a generalizable approach for linking deep learning to pathophysiological processes in human disease.

neuroscience↗

bioRxiv2018

Effect of continuous compressive force on the expression of RANKL, OPG, and VEGF in MC3T3-E1 and MLO-Y4 cells

Osteocytes, known to have mechano-sensory functions, influence the regulation of bone remodeling. However, the mechanism by which osteocytes regulate bone metabolism when mechanical forces are being applied is still unclear. Osteoclastogenesis is mainly regulated by receptor activator of nuclear factor kappa-B ligand (RANKL); the protein osteoprotegerin (OPG) and angiogenesis also play important roles in osteogenesis. RANKL, OPG, and vascular endothelial growth factor (VEGF) are thought to be key factors for bone metabolism. In this study, we examined the effect of a continuous compressive force (CF) on the expression of RANKL, OPG, and VEGF in osteoblastic murine osteocytes (MLO-Y4) and osteoblastic (MC3T3-E1) cells. Gene and protein expression levels of RANKL, OPG, and VEGF in MLO-Y4 and MC3T3-E1 cells were quantitatively determined by real-time PCR and enzyme-linked immunosorbent assay (ELISA). Both cell types were also subjected to a CF of 1.0 g/cm2 for 1, 3, 6, and 12 hours. Furthermore, the effect of a stretch-activated (S-A) channel was examined by gadolinium (Gd3+) administration. The ratio of gene and protein expressions of RANKL, VEGF, and RANKL/OPG in MLO-Y4 cells were significantly higher than in MC3T3-E1 cells, while the expression of OPG was significantly lower. After CF application, both cell types showed significant increases in RANKL and VEGF expression as well as the RANKL/OPG ratio. Additionally, the upregulated gene and protein levels of these factors were reduced by Gd3+ administration.\n\nThese findings suggest that osteocytes play more important roles in bone metabolism and angiogenesis than osteoblasts. Osteocytes regulate the expression of RANKL, OPG, and VEGF via the S-A channel through the response to mechanical stress.

cell biology↗