Biology subjects

Barai, P.

Publications and source records attributed to Barai, P..

3 recordsLinked to original sources

bioRxiv2026

Loss of LanC-like proteins impairs post-injury regeneration of aging muscles

The adult skeletal muscle regenerates robustly upon injury, but this regenerative capacity rapidly declines with age. In this study, we identify the lanthionine synthetase C-Like (LanCL) proteins, mammalian homologs of the bacterial peptide cyclase LanC, as positive regulators of muscle regeneration in middle-aged mice. In a barium chloride-induced injury model, we found the protein levels of LanCL1 and LanCL2 to increase during an early phase of regeneration in middle-aged (12-month-old) but not young adult (4-month-old) mice. Utilizing a mouse line lacking all three LanCL proteins (LanCL triple KO or LTKO), we examined a potential role of LanCL in injury-induced muscle regeneration. Consistent with an age-dependent function of LanCL, we observed a delayed regeneration of the tibialis anterior (TA) muscle after injury, as reflected by reduced sizes of regenerating myofibers in middle-aged (but not young) LTKO compared to age-matched WT mice. Although the pool size of quiescent satellite cells (Pax7+) was comparable between 12-month-old LTKO and WT muscles without injury, the number of Pax7+ cells was significantly higher in regenerating LTKO muscles at day 5 after injury, accompanied by drastically decreased numbers of MyoD+ and MyoG+ cells, as well as increased numbers of proliferating cells. In addition, we detected elevated expression of pro-inflammatory cytokines in regenerating LTKO muscles, while the number of macrophages was similar comparing LTKO and WT muscles. Taken together, our observations suggest that in aging muscles LanCLs are important for proper timing of inflammation resolution and regeneration upon injury.

physiology↗

bioRxiv2024

Injury-induced transcription in the planarian outer epithelium is critical for tissue regeneration

Planarian flatworms have an extraordinary regenerative capacity; even a small, asymmetrical fragment of amputated planarian tissue can recreate an entirely new animal. The planarian field has made significant progress in identifying specific genes and cell types required for this complex process, but substantially less is known about the molecular mechanisms that convert a significant physical injury (e.g., head amputation) into the expression of specific genes in particular cell types. One tissue in which this question is particularly relevant is the planarian epidermis, a single-layer, mucociliary epithelium with similarities to the epithelia of mammalian airways. This epithelium plays an essential, early role in planarian regeneration as the cells surrounding the wound site must quickly stretch and extend to cover the wound area after injury. We hypothesized that these injury-induced morphological changes activate the transcription of genes with essential functions in the regenerative process. To best detect these transcriptional changes, we developed a rapid method for isolating the planarian outer epithelium and prepared ribodepleted RNA-sequencing libraries from samples isolated at multiple time points after tissue amputation. One gene we both identified using these methods and found to be functionally important for regeneration is a putative planarian homolog of vertebrate Shoc2. SHOC2 is an essential scaffolding protein that mediates specific, context-dependent activation of the ERK1/2 signaling pathway (J O_SCPLOWANGC_SCPLOW O_SCPLOWANDC_SCPLOWG O_SCPLOWALPERINC_SCPLOW2016). Notably, the ERK1/2 pathway is known to be activated after injury and required for regeneration in multiple species (M O_SCPLOWANUELC_SCPLOW et al. 2006; O O_SCPLOWWLARNC_SCPLOWet al. 2017; T O_SCPLOWOMASSOC_SCPLOWet al. 2023). These findings suggest that these epithelial datasets have the potential to uncover many functionally relevant and possibly highly conserved genes that play fundamental roles in animal regeneration.

developmental biology↗

bioRxiv2024

RNaseH-based ribodepletion of total planarian RNA improves detection of longer and non-polyadenylated transcripts

The overwhelming majority of RNA species isolated from cells or tissues using organic extraction are ribosomal RNAs (rRNA), whereas a relatively small percentage are messenger RNAs (mRNA). For studies that seek to detect mRNA transcripts and measure changes in their expression, this lopsided ratio of desired transcripts to undesired transcripts creates a significant challenge to obtaining sensitive and reproducible results. One method for improving mRNA detection is to selectively amplify polyadenylated (polyA) mRNA molecules when generating RNA-seq libraries, a strategy that is generally very successful in many species. However, this strategy is less effective when starting with total RNA from some species e.g., the planarian species Schmidtea mediterranea (S.med), as it generates libraries that still contain significant and variable amounts of rRNA reads. Further, commercially available ribodepletion kits do not efficiently deplete rRNAs from these samples because their sequences are divergent from mammalian rRNAs. Here we report a customized, optimized, and economical ribodepletion strategy than allows the generation of comprehensive RNA-seq libraries with less than one percent rRNA contamination. We show that this method improves transcript detection, particularly for those without polyA tails (e.g., core histones) and those that are relatively long (e.g., microtubule motor proteins). Using this custom ribodepletion approach, we also detected many transcripts that are not represented in the most recent set of S.med gene annotations, including a subset that are likely expressed transposable elements (TEs). To facilitate future differential expression analyses of these newly identified loci, we created both an annotation file of the new loci we identified and a bioinformatic pipeline for generating additional annotations from future libraries. As significant recent research shows that TE activation is regulated and functionally important, the resources provided here will provide a starting point for investigating such mechanisms in planarians and other species with less conserved rRNA sequences.

molecular biology↗