Molecular and Cellular Biology最新文献

Copper is an essential but potentially toxic nutrient required for a variety of biological functions. Mammalian cells use a complex network of copper transporters and metallochaperones to maintain copper homeostasis. Previous work investigating the role of copper in various disease states has highlighted the importance of copper transporters and metallochaperones. However, questions remain about how copper distribution changes under dynamic conditions like tissue differentiation. We previously reported that the copper exporter ATP7A is required for skeletal myoblast differentiation and that its expression changes in a differentiation dependent manner. Here, we sought to further understand the ATP7A-mediated copper export pathway by examining ATOX1, the copper chaperone that delivers copper to ATP7A. To investigate the role of ATOX1 in a dynamic cellular context, we characterized its protein-protein interactions during myoblast differentiation using the proximity labeling protein APEX2 to biotinylate proteins near ATOX1. We discovered that the ATOX1 interactome undergoes dramatic changes as myoblasts differentiate. These dynamic interactions correlate with distinct phenotypes of ATOX1 deficiency in proliferating and differentiated cells. Together, our results highlight the dynamic interactome of ATOX1 and its contribution to myoblast differentiation.

Transcription factors (TFs) are traditionally classified as activators or repressors, yet some can perform both roles. We highlight well-supported examples of dual activator/repressor functions and review the mechanisms that explain how duality arises. These examples reveal that transcriptional duality arises from three recurring mechanisms: positional effects, cofactor exchange, and regulatory switches. Even within these recurring mechanisms, the precise molecular details diverge, with regulatory outcomes dictated by differences in TF positioning, cofactor availability, modification state, and ligand binding. We propose that future work should move beyond descriptive labels of context specificity and instead focus on elucidating the precise molecular mechanisms by which TFs function to elicit opposing regulatory effects.

Cellular senescence has a dual role in both tumor suppression and the promotion of age-related diseases. This paradox suggests the existence of functionally distinct "beneficial" and "detrimental" senescent states, yet the molecular basis that governs their fate has remained elusive. Here, we reveal that the dynamic exchange of histone H2AX on chromatin functions as an essential quality control mechanism that dictates the quality of senescence. We demonstrate that the histone acetyltransferase TIP60, in complex with the chaperone FACT, acetylates H2AX at lysine 5 (K5), which in turn drives its dynamic exchange. This histone exchange is indispensable for promoting the degradation of the DNA damage response mediator MDC1, a process we uncover is mediated by a novel DNA-PKcs-p97 signaling axis. Disruption of this TIP60-FACT-H2AX exchange pathway leads to the hyperaccumulation of MDC1 and a shift toward error-prone nonhomologous end joining (NHEJ), inducing a pathological senescent state with oncogenic potential. Our study redefines histone exchange from a passive chromatin event to an active regulatory hub that determines the fate of aging cells. These findings provide a molecular basis for the heterogeneity of senescence and establish a rationale for developing "senomorphic" therapies aimed at improving the quality of aging.

The brain is one of the most lipid-rich organs, reflecting the critical role of lipid metabolism in neuronal and glial cell function. While mitochondria are central to energy metabolism, calcium signaling, and cell death, they do not utilize lipid oxidation for energy but rely on lipids for membrane integrity and intracellular communication. Here we review the interactions between lipids and mitochondria in intracellular signaling within brain cells, examining their roles in normal physiology and the mechanisms underlying major neurodegenerative diseases. Alterations in lipid homeostasis and mitochondrial metabolism are implicated in neurodegeneration, highlighting the importance of lipid-mediated mitochondrial signaling pathways. Understanding these interactions provides insights into cellular dysfunction in neurodegenerative disorders and may inform future therapeutic strategies targeting lipid and mitochondrial pathways.

Crohn's disease (CD) is an inflammatory gastrointestinal disorder marked by impaired autophagy due to inefficient bacterial uptake. We studied the effects of autophagy modulation using Tat-beclin-1 and carbamazepine (CBZ) on dendritic cells (DCs) and Paneth cell functionality in pediatric CD patients. Twenty CD children genotyped for the ATG16L1 rs2241880 polymorphism and 10 healthy controls were enrolled. DCs were incubated with fluorochrome-conjugated particles of Escherichia coli or DQ-ovalbumin after pretreatment with CBZ or Tat-beclin-1 to evaluate antigen processing. Treated DCs were stained for P62, LAMP1, and LC3, and analyzed by confocal microscopy. Paneth cells from biopsies were pretreated with both drugs, stained for lysozyme, and analyzed by transmission electron microscopy. Antigen processing increased after Tat-beclin-1 and CBZ treatment in all groups. DCs expressed higher activation markers HLA-DR and CD86+, notably in high-risk patients, who also showed increased DQ-OVA processing. The number of lysozymes in Paneth cells from controls did not change after Tat-beclin-1 treatment, while in the CD group, it decreased significantly, suggesting increased exocytosis. CBZ treatment increased secretory granules only in CD inflamed tissue. Our results indicate that CBZ and Tat-beclin-1 enhance autophagic flux, representing a novel approach to treating pediatric CD patients.

p53 is a key tumor suppressor, and mutations in the p53 gene occur in more than half of all human cancers. p53, which is under tight and complex regulation in cells, functions primarily as a transcription factor regulating genes involved in many cellular processes, including cell cycle arrest, apoptosis, senescence, ferroptosis, and metabolism, thereby maintaining genomic integrity and preventing tumorigenesis. While the cell-intrinsic functions of p53, which contribute to its tumor-suppressive activity, have been extensively studied, it is now clear that p53 also plays an important role in immune regulation, a connection first observed when p53 was identified as a cellular protein interacting with viral antigens. Growing evidence shows that p53 modulates both innate and adaptive immunity by regulating cytokine production, antigen presentation, and the functions of immune cells, thereby contributing to host defense against infections, inflammatory responses, and antitumor immunity. In this review, we summarize and discuss the multifaceted roles of p53 and its signaling in regulating immune functions and their implications in human diseases, particularly cancer. A better understanding of the immune-related functions of p53 is crucial for advancing cancer treatment and broadening insights into immunity and disease.

The telomerase RNA-protein enzyme is critical for most eukaryotes to complete genome copying by extending chromosome ends, thus solving the end-replication problem and postponing senescence. Despite the importance of the fission yeast Schizosaccharomyces pombe to biomedical research, very little is known about the structure of its 1212 nt telomerase RNA. We have determined the secondary structure of this large RNA, TER1, based on phylogenetics and bioinformatic modeling, as well as genetic and biochemical analyses. We find several conserved regions of the rapidly evolving TER1 RNA are important to maintain telomeres, based on testing truncation mutants in vivo, whereas many other large regions are dispensable. This is similar to budding yeast telomerase RNA, and consistent with functioning as a flexible scaffold for the RNP. We tested if the essential three-way junction works from other locations in TER1, finding that it can, supporting that it is flexibly scaffolded. Furthermore, we find that a half-sized Mini-TER1 allele, built from the catalytic core and the three-way junction, reconstitutes catalytic activity with TERT in vitro. Overall, we provide a secondary structure model for the large fission-yeast telomerase lncRNA, based on phylogenetics and molecular-genetic testing in cells, and insight into the RNP's physical and functional organization.

The serine/threonine kinase, TOR (target of rapamycin), exists in two complexes, namely TORC1 (with either Tor1 or Tor2 kinase) and TORC2 (that contains Tor2, but not Tor1), and its pharmacological inhibition by rapamycin impairs the PIC (pre-initiation complex) formation at the ribosomal protein genes (and hence transcription and ribosome biogenesis). However, TOR's involvement in such gene regulation has not been elucidated genetically at the level of Tor1, Tor2, TORC1 or TORC2. Here, we demonstrate that null mutation of TOR1 and short-term depletion of its expression do not affect the PIC formation (and transcription) at the ribosomal protein genes. Likewise, PIC formation and transcription are not altered in TORC2-specific tor2-tsA conditional mutant or following short-term depletion of TOR2 expression. These results support the dispensability of TORC2 for ribosomal protein gene expression, and indicate that Tor1 and Tor2 play redundant roles via TORC1 for PIC formation, and hence transcription. In agreement, the Δtor1 mutant in combination with both TORC1 and TORC2-specific tor2-tsC conditional mutation impairs PIC formation at the ribosomal protein genes with consequent reduction in transcription. Collectively, our genetic analysis support redundant, yet distinct, functions of Tor1 and Tor2 via TORC1, not TORC2, in regulation of the ribosomal protein gene expression.

A-to-I RNA editing introduces A-to-G variation at post-transcriptional level, but it remains mysterious. What is the advantage of functional RNA editing compared to an A/G heterozygous SNP? Here, we provide the following situations that particular RNA editing sites can be superior to heterozygous SNPs even independent of its temporospatial regulation. (1) Assume a site with A/G heterozygote advantage. RNA editing does not undergo Mendelian segregation and recombination that inevitably produce homozygotes of lower fitness. (2) Graded RNA editing level. A snapshot of editing profile shows strong tissue-specific editing levels, providing flexible stoichiometry of edited/unedited versions, while heterozygous SNPs generally produce similar expression of two alleles. (3) Higher molecular diversity. N RNA editing sites in a gene theoretically produce a dramatic number of X = 2^N mRNA haplotypes, but all SNPs in a gene can only produce two alleles. Nevertheless, we emphasize that these advantageous sites may emerge through complicated evolutionary process and remain rare across the genome. We systematically discussed the pros and cons of RNA editing versus heterozygous SNPs, deepening our understanding of the biological functions of cis-regulatory mechanisms. We provide putative answers to why evolution chose RNA editing instead of a genomic mutation at particular sites.

DNA methylation inhibitors are widely used in treating myeloid malignancies, yet their precise effects on chromatin organization and nuclear architecture remain incompletely understood. Here, the integrated molecular, cellular, and biophysical approaches to investigate the impact of azacitidine (AZA) and decitabine (DEC) on chromatin structure and nuclear mechanics in AML-007 leukemia cells are presented. Confocal microscopy revealed drug-induced alterations in nuclear morphology and actin cytoskeleton organization, with DEC inducing significant nuclear enlargement and disorganization at lower concentrations (1.0 µM) compared to AZA (5.0 µM). Chromatin condensation assays demonstrated that DEC increased chromatin accessibility in a concentration-dependent manner, while AZA produced subtler effects. Optical tweezers measurements showed both agents reduced nuclear stiffness, with DEC exerting a greater impact. Spectroscopic analysis confirmed differential drug incorporation into DNA, with higher methylation loss and structural changes observed following DEC treatment. Refractive index mapping revealed chromatin decompaction, aligning with increased accessibility and nuclear softening. These findings demonstrate that DNA hypomethylating agents exert distinct, concentration-dependent effects on nuclear organization and chromatin structure, which can be quantified through molecular and biophysical readouts. This study underscores the value of integrative methods for revealing epigenetic drug effects on chromatin architecture in leukemia cells.