Reimer, B. M. | Search bioRxiv

bioRxiv2025

A frameshift mutation in the methyltransferase rlmN is associated with increased linezolid resistance in Mycobacterium tuberculosis

Linezolid is a key component of treatment regimens for multidrug-resistant and extensively drug-resistant tuberculosis, which is caused by the pathogen Mycobacterium tuberculosis (MTB). Resistance to linezolid in MTB has traditionally been attributed to mutations in the 23S rRNA (rrl) and ribosomal protein L3 (rplC), but only a fraction of clinically observed linezolid resistance is explained by mutations in these two genes. We report that an analysis of strains with paired whole-genome sequencing and linezolid minimum inhibitory concentration (MIC) phenotyping from the Bacterial and Viral Bioinformatics Resource Center (BV-BRC) reveals that a relatively common frameshift mutation in MTB methyltransferase rlmN (5.3% of assessed isolates) is significantly associated with increased linezolid MIC. In additional to statistical associations, we provide evolutionary evidence of homology to an established linezolid resistance mechanism in Staphylococcus aureus, and structural evidence that the frameshift mutation likely ablates rlmN methyltransferase functionality.

bioinformatics↗

bioRxiv2025

Mechanistic evidence that motif-gated domain recognition drives contact prediction in protein language models

Protein language models (pLMs) achieve state-of-the-art performance on protein structure and function prediction tasks, yet their internal computations re-main opaque. Sparse autoencoders (SAEs) have been used to recover sparse features, called latents, from pLM layer representations, whose activations cor-relate with known biological concepts. However, prior work has not established which model concepts are causally necessary for pLM performance on down-stream tasks. Here, we adapt causal activation patching to the pLM setting and perform it in SAE latent space to extract the minimal circuit responsible for accuracy in a contact prediction task for two case study proteins. We observe that preserving only a tiny fraction of latent-token pairs (0.022% and 0.015%) is sufficient to retain contact prediction accuracy in a residue unmasking experiment. Our circuit indicates a two-step computation in which early-layer motif detectors respond to short local sequence patterns, gating mid-to-late domain detectors which are selective for protein domains and families. Path-level ab-lations confirm the causal dependence of domain latents on upstream motif latents. To evaluate these components quantitatively, we introduce two diagnostics: a Motif Conservation Test and a Domain Selectivity Framework that supports hypothesis-driven tests. All candidate motif-detector latents pass the conservation test, and 18/23 candidate domain-detector latents achieve AUROC [≥]0.95. To our knowledge, this is the first circuits-style causal analysis for pLMs, pin-pointing the motifs, domains, and motif-domain interactions that drive contact prediction in two specific case studies. The framework introduced herein will enable future mechanistic dissection of protein language models. Code available at https://github.com/NainaniJatinZ/plm_circuits

bioinformatics↗