Biology subjects

Rodriguez, A. I.

Publications and source records attributed to Rodriguez, A. I..

3 recordsLinked to original sources

bioRxiv2025

Non-redundant cardiolipin synthases support membrane integrity and stress resilience in Bacteroides fragilis

Gut-resident bacteria must tolerate diverse membrane-disrupting agents, including bile acids, to maintain colonization. Cardiolipin is an anionic phospholipid that supports membrane integrity and stress resilience in many bacteria. However, cardiolipin synthases remain poorly characterized in the Bacteroidota, a dominant phylum of the human gut microbiota. The prevalent gut commensal Bacteroides fragilis encodes two predicted cardiolipin synthases, ClsA and ClsB. We previously identified both cls genes as bile-acid fitness factors in B. fragilis P207, but the individual contributions of ClsA and ClsB to cell physiology had remained undefined. Here we combine targeted gene deletion with high-resolution lipidomics, metabolomics, and elemental mass spectrometry to show that the two enzymes have non-redundant functions in the cell. Cardiolipin is a minor lipid in the B. fragilis membrane, and both Cls enzymes contribute to its production. clsA and clsB differ in growth-phase expression, in their effects on cell morphology, and in their associated cardiolipin species. Loss of each enzyme also produces distinct changes in fitness under several membrane-perturbing stresses and in the broader cellular metabolome, including compound classes with documented bioactivity in mammalian hosts. In contrast to the acute ion-gradient disruption caused by the secondary bile acid deoxycholate, deletion of both synthases did not measurably alter steady-state intracellular ion levels under standard growth conditions, indicating that cardiolipin loss does not perturb basal ion homeostasis under these conditions. Together, these results define non-redundant roles for two cardiolipin synthases in a common member of the human gut microbiota. ImportanceInflammatory bowel diseases affect millions of people worldwide. The gut bacterium Bacteroides fragilis is a normal member of the human intestinal microbiota that can also bloom to high abundance in inflamed guts. To survive within the gut, B. fragilis must maintain the integrity and function of its cell membrane. In this study, we characterize the functional role of two B. fragilis genes that contribute to the synthesis of the membrane lipid, cardiolipin. We find that the two cardiolipin synthase genes are not functionally interchangeable; each impacts the cells lipid pool in distinct ways and contributes differently to how B. fragilis responds to membrane stress. Our work provides insight into how a common gut bacterium adapts to conditions encountered in the intestine, and improves understanding of membrane biology in this important group of gut microbes.

microbiology↗

bioRxiv2025

Coarse-grained chromatin dynamics by trackingmultiple similarly labeled gene loci

The "holy grail" of chromatin research would be to follow the chromatin configuration in individual live cells over time. One way to achieve this goal would be to track the positions of multiple loci arranged along the chromatin polymer with fluorescent labels. Use of distinguishable labels would define each locus uniquely in a microscopic image but would restrict the number of loci that could be observed simultaneously, because of experimental limits to the number of distinguishable labels. Use of the same label for all loci circumvents this limitation but requires a (currently lacking) framework for how to establish each observed locus identity, i.e. to which genomic position it corresponds. Here we analyze theoretically, using simulations of Rouse-model polymers, how single-particle-tracking of multiple identically-labeled loci enables determination of loci identity. We show that the probability of correctly assigning observed loci to genomic positions converges exponentially to unity as the number of observed loci configurations increases. The convergence rate depends only weakly on the number of labeled loci, so that even large numbers of loci can be identified with high fidelity by tracking them across about 8 independent chromatin configurations. In the case of two distinct labels that alternate along the chromatin polymer, we find that the probability of the correct assignment converges faster than for same-labeled loci, requiring observation of fewer independent chromatin configurations to establish loci identities. Finally, for a modified Rouse-model polymer, that realizes a population of dynamic loops, we find that the success probability also converges to unity exponentially as the number of observed loci configurations increases, albeit slightly more slowly than for a classical Rouse model polymer. Altogether, these results establish particle tracking of multiple identically- or alternately-labeled loci over time as a feasible way to infer temporal dynamics of the coarse-grained configuration of the chromatin polymer in individual living cells. SIGNIFICANCEIn spite of recent success in elucidating its spatial organization, chromatins time-dependent, dynamical behavior remains far less studied, and correspondingly much less understood. To address the critical need to elucidate chromatin dynamics, this paper proffers a route towards an experimental characterization of coarse-grained chromosomal dynamics, via particle tracking of multiple labeled loci, labeled with just one or two different fluophor colors or intensities. Theoretically, we show that particle tracking of multiple identically labeled loci across only about 8 independent chromatin configurations should be a feasible way to establish the time-dependent, coarse-grained configuration of the chromatin polymer in individual living cells.

biophysics↗

bioRxiv2025

ER-associated degradation relying on protein O-mannosylation

Protein quality control in the secretory pathway, whose dysfunction is linked to various human diseases, begins in the endoplasmic reticulum (ER) and involves ER-associated degradation (ERAD) of terminally misfolded species. We have addressed the roles of protein O-mannosylation in ERAD using complementary genome-wide screens in yeast. Our findings reveal that protein O-mannosylation by the conserved ER-resident O-mannosyltransferase complex Pbn1-Gpi14 generates ERAD cues. This mechanism participates in regulating ERAD of substrates that contain serine-rich regions and provides a fail-safe mechanism for the degradation of non-N-glycosylated misfolded proteins. Our data suggest that the de novo synthesis of ERAD cues through protein O-mannosylation is an essential component of ER quality control.

cell biology↗