Biology subjects

Rahlwes, K. C.

Publications and source records attributed to Rahlwes, K. C..

6 recordsLinked to original sources

bioRxiv2026

Sorting nexin 5 mediates antigen presentation and immunity against Mycobacterium tuberculosis

Tuberculosis (TB) remains one of the leading causes of death from a single infectious agent worldwide, yet the host pathways that regulate antigen presentation and lung inflammation during Mycobacterium tuberculosis (Mtb) infection are incompletely defined. Sorting nexin 5 (SNX5) is a protein best known for roles in endosomal trafficking, antigen processing, and antiviral host defense, but its contribution to immunity during Mtb infection is unknown. Here, we show that SNX5-deficient mice exhibit markedly increased mortality following low-dose aerosol infection despite unchanged pulmonary bacterial burden compared to wild-type mice. Snx5-/- mice developed exacerbated lung inflammation without major alterations in immune cell recruitment. In macrophages, Snx5 did not affect phagocytosis, vacuolar maturation, intracellular bacterial control, or global transcriptional responses to Mtb, but was required for efficient MHC class II antigen presentation. Snx5 deficiency reduced antigen degradation, limited peptide loading onto MHC-II and impaired activation of antigen-specific CD4+ T cells without altering surface MHC-II abundance or expression of costimulatory molecules. Together, these findings identify SNX5 as a previously unrecognized regulator of MHC-II peptide loading that shapes inflammatory outcomes during pulmonary Mtb infection, highlighting a role for the endosomal sorting machinery in immunity to intracellular pathogens.

immunology↗

bioRxiv2025

SMURF2 inhibits autophagic control of Mycobacterium tuberculosis in macrophages

Autophagy is a critical host defense mechanism that restricts intracellular pathogens such as Mycobacterium tuberculosis (Mtb). A key step in this process is the ubiquitination of Mtb or Mtb-associated structures. The E3 ligase SMURF1 catalyzes K48-linked ubiquitination, promoting bacterial clearance. However, the function of its homolog, SMURF2, in host defense remains undefined. Here, we demonstrate that Smurf2 deletion in murine macrophages increases SMURF1 levels, enhances LC3B lipidation, augments K48 ubiquitination of Mtb-associated structures, and reduces intracellular Mtb replication. These effects are reversed by Smurf1 deletion, indicating that SMURF2 restricts autophagy in a SMURF1-dependent manner. Mice with myeloid-specific Smurf2 deletion exhibit modestly prolonged survival following aerosol Mtb infection. In human macrophages, SMURF2 knockdown or its pharmacological inhibition with the HECT ligase inhibitor Heclin reduces Mtb replication. Together, our findings identify SMURF2 as a negative regulator of selective autophagy and host immunity to Mtb and suggest that targeting SMURF2 may represent a novel host-directed therapeutic strategy for tuberculosis.

immunology↗

bioRxiv2025

Deubiquitinase USP15 restricts autophagy and macrophage immunity to Mycobacterium tuberculosis

Autophagy enables macrophages to degrade intracellular Mycobacterium tuberculosis (Mtb), and this defense depends on E3 ubiquitin ligases such as PARKIN and SMURF1, which tag Mtb-associated structures for lysosomal clearance. Deubiquitinases (DUBs) counter ubiquitin ligases by removing ubiquitin from molecular targets. We hypothesized that DUBs might offset ubiquitin ligase activity and negatively regulate host immunity to Mtb. Here, we identify ubiquitin-specific protease 15 (USP15) as a negative regulator of autophagy-mediated macrophage immunity to Mtb. Using a targeted knockdown screen in mouse macrophages, we found that Usp15 loss increased K63-linked ubiquitination and LC3 recruitment to Mtb-associated structures, leading to reduced bacterial replication. These effects required USP15s catalytic activity and were reversed by knockdown of PARKIN (Park2) or inhibition of autophagy initiation. In primary human macrophages, USP15 knockdown similarly enhanced LC3 targeting and restricted Mtb growth. Importantly, pharmacologic inhibition of USP15 with a selective small molecule decreased Mtb burden in human macrophages. Our findings identify USP15 as a suppressor of macrophage immunity and suggest that targeting deubiquitinases may represent a promising host-directed therapeutic strategy against tuberculosis.

immunology↗

bioRxiv2025

Mycobacterial Phenolic Glycolipid Triggers ATP-Mediated Neuronal P2X3 Signaling and Cough

Cough drives respiratory pathogen transmission, yet how microbes directly engage host sensory neurons to trigger cough is largely unknown. We previously demonstrated that the Mycobacterium tuberculosis (Mtb) glycolipid sulfolipid-1 (SL-1) activates neurons and induces cough. Here, we reveal that phenolic glycolipid (PGL) produced by the hypertransmissible HN878 Mtb strain activates both mouse and human nociceptive neurons in vitro using calcium imaging and electrophysiology and is sufficient to induce cough using plethysmography. Combined with SL-1, PGL potently triggers neuronal activation. By synthesizing various PGL analogs, we show that neuroactivity is proportional to saccharide chain length and structure. Mechanistically, PGL stimulates rapid extracellular ATP release, which engages neuronal P2X3 purinergic receptors--an effect blocked by a P2X3 antagonist. These findings uncover a neuronal activation pathway co-opted by certain Mtb strains to enhance transmission via cough and suggest inhibition of purinergic signaling as a potential strategy to block airborne spread of Mtb.

neuroscience↗

bioRxiv2020

The role of LmeA, a mycobacterial periplasmic protein, in stabilizing the mannosyltransferase MptA and its product lipomannan under stress

The mycobacterial cell envelope has a diderm structure, composed of an outer mycomembrane, an arabinogalactan-peptidoglycan cell wall, periplasm and an inner membrane. Lipomannan (LM) and lipoarabinomannan (LAM) are structural and immunomodulatory components of this cell envelope. LM/LAM biosynthesis involves a number of mannosyltransferases and acyltransferases, and MptA is an 1,6-mannosyltransferase involved in the final extension of the mannan backbones. Recently, we reported the periplasmic protein LmeA being involved in the maturation of the mannan backbone in Mycobacterium smegmatis. Here, we examined the role of LmeA under stress conditions. We found that the lmeA transcription was upregulated under two stress conditions: stationary growth phase and nutrient starvation. Under both conditions, LAM was decreased, but LM was relatively stable, suggesting that maintaining the cellular level of LM under stress is important. Surprisingly, the protein levels of MptA were decreased in lmeA deletion mutant ({Delta}lmeA) in both stress conditions. The transcript levels of mptA in {Delta}lmeA were similar to or even higher than those in the wildtype, indicating that the decrease of MptA protein was a post-transcriptional event. Consistent with the decrease in MptA, {Delta}lmeA was unable to maintain the cellular level of LM under stress. Even during active growth, overexpression of LmeA led the cells to produce more LM and become more resistant to several antibiotics. Altogether, our study reveals the roles of LmeA in the homeostasis of the MptA mannosyltransferase particularly under stress conditions, ensuring the stable expression of LM and the maintenance of cell envelope integrity.

microbiology↗

bioRxiv2019

Discovery of an essential mycobacterial polar growth factor involved in cell envelope integrity and LM/LAM transport

Mycobacteria, including the human pathogen Mycobacterium tuberculosis, grow by inserting new cell wall material at their poles. This process and that of division are asymmetric, producing a phenotypically heterogeneous population of cells that respond non-uniformly to stress (Aldridge et al., 2012; Rego et al., 2017; Richardson et al., 2016). Surprisingly, deletion of a single gene - lamA - leads to more symmetry, and to a population of cells that is more uniformly killed by antibiotics (Rego et al., 2017). How does LamA create asymmetry? Here, using a combination of quantitative time-lapse imaging, bacterial genetics, and lipid profiling, we find that LamA recruits essential proteins involved in cell wall synthesis to one side of the cell - the old pole. One of these proteins, MSMEG_0317, here renamed PgfA, was of unknown function. We show that PgfA is a periplasmic protein that interacts with MmpL3, an essential transporter that flips mycolic acids in the form of trehalose monomycolate (TMM), across the plasma membrane. PgfA interacts with a TMM analog suggesting a direct role in TMM transport. Yet our data point to a broader function as well, as cells with altered PgfA levels have differences in the abundance of other lipids and are differentially reliant on those lipids for survival. Overexpression of PgfA, but not MmpL3, restores growth at the old poles in cells missing lamA. Together, our results suggest that PgfA is a key regulator of polar growth and cell envelope composition in mycobacteria, and that the LamA-mediated recruitment of this protein to one side of the cell is a required step in the establishment of cellular asymmetry.

microbiology↗