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Expression of tardigrade intrinsically disordered proteins in C. elegans reveals variable effects on aging and stress

Anna Entin, Daniel HicksiD, Elijah A. Roditi, Juri KimiD, Aeowynn J. CoakleyiD, Adam J. HrubyiD, Athena AlcalaiD, Naibedya Dutta, Maxim AverbukhiD, Shivay ParihariD, Rebecca Aviles Barahona, Thomas Jiyoung Lee, Veronica Kuo, Vinaya SahasrabuddheiD, Bharat Vivan ThapaiD, Jamie Dale KraftiD, Peter J. MulleniD, Gilberto GarciaiD, Ryo Higuchi‐SanabriaiD

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In-depth summary

Claude Opus 5.5 · xhigh effort · 8 Nov 2026 · from the full text of version 1 (31 Oct 2026) · CC BY 4.0

Constitutive somatic expression of four tardigrade disordered proteins shortened C. elegans lifespan and healthspan and gave little stress protection. Three of the four reduced polyQ aggregation under hyperosmotic stress.

Germline expression of the tardigrade protein DSUP was reported to improve stress resistance and lifespan in C. elegans, but somatic tissues, the usual therapeutic target, differ from germ cells. This paper tests CAHS, MAHS, DSUP and RvLEAM expressed in somatic tissues of whole animals.

Key results

  1. ShownSomatic DSUP expression (sur-5p) shortened lifespan and reduced thrashing and brood size. Intestine or hypodermis expression shortened lifespan, but muscle or neuron expression did not.

    Lifespan was significantly reduced, including on kanamycin-killed bacteria. Thrashing was lower at all time points. RNAi timing with dcr-1 showed development-only expression was sufficient to cut lifespan, while adult-only expression had a mild but significant effect. · Somatic DSUP expression exhibits toxic effects

  2. ShownMAHS, CAHS and RvLEAM also shortened lifespan and reduced thrashing and brood size, so the toxicity is shared across TDP classes. The MAHS lifespan effect disappeared on kanamycin-killed bacteria.

    Significant reductions in lifespan, in thrashing at all time points, and in brood size for all three. CAHS and RvLEAM still shortened lifespan on kanamycin-killed bacteria. · Expression of TDPs in C. elegans broadly reduces…

  3. ShownStress protection was narrow. DSUP lowered survival after heat and UVC and nothing changed under oxidative stress. The other three TDPs did not alter heat, UV or oxidative survival. DSUP, CAHS and RvLEAM, but not MAHS, reduced polyQ foci under 500 mM NaCl.

    Log-rank tests for heat, Mann-Whitney tests for UVC and polyQ foci. Heat-inducible hsp-16.2 expression of CAHS or MAHS did not improve thermotolerance. · Somatic DSUP expression exhibits toxic effects; Expression of TDPs in C. elegans broadly reduces…; Lifespan and thermotolerance measurements; Figure legends

  4. SuggestedNo evidence of mitochondrial dysfunction as the primary driver of the phenotypes: basal oxygen consumption was unchanged for all four TDPs, and mitochondrial morphology changed only in isolated tissue and age combinations.

    Seahorse respirometry showed no significant change. mitoMAPR found fewer branch points with CAHS in day 7 hypodermis and shorter mitochondria with MAHS in day 1 intestine. Muscle was unaffected, and UPRmt and oxidative-stress gene sets were not coordinately induced. · Expression of TDPs in C. elegans broadly reduces…; Seahorse analysis

  5. ShownEach TDP produced a largely distinct transcriptional response rather than a shared program.

    Bulk RNA-seq (three replicates) gave 986 DEGs for RvLEAM, 692 for MAHS, 650 for DSUP and 74 for CAHS. Only 22 DEGs were shared by all four. GO enrichment showed overlapping themes (immune defense, cuticle) with TDP-specific gene patterns, and RvLEAM alone enriched ER stress terms. · Expression of TDPs in C. elegans broadly reduces…; RNA-seq

Methods, data and reproducibility

Transgenic C. elegans lines expressed codon-optimized R. varieornatus CAHS, MAHS, DSUP and RvLEAM from tissue-specific promoters, then underwent lifespan, thrashing, brood, stress, polyQ aggregation, respirometry, mitochondrial imaging and bulk RNA-seq. RNAi against dsup, combined with dcr-1 RNAi, switched expression on or off at defined stages.

  • Transgenic TDP-expressing lines (sur-5p, col-19p, myo-3p, rgef-1p, vha-6p) versus N2 controls; lifespan, thrashing, brood and mitochondrial imaging; per-assay n reported in figures and Table S2 · C. elegans
  • Oxidative stress survival (6 mM tert-butyl hydroxide), independent biological replicates · C. elegans · n = N = 77-101 (DSUP); N = 74-97 (RvLEAM, CAHS, MAHS)
  • Bulk RNA-seq of L4 animals for four TDP strains and N2 · C. elegans · n = three biological replicates per condition · PRJNA1550449
Methods & tools
GraphPad Prism 10, mitoMAPR, Seahorse XFe96 Analyzer, Trim Galore v0.6.7-1, STAR-2.7.3a, Subread v2.0.3, DESeq2, sva v3.54.0, WormEnrichr, Illumina NovaSeq6000 (Novogene), WBcel235 genome
Software & environment
Software versions given for Trim Galore (v0.6.7-1), STAR (2.7.3a), Subread (v2.0.3) and sva (v3.54.0); no container or requirements file stated.
Reproducibility gaps
Raw RNA-seq is deposited in NCBI SRA (PRJNA1550449). Per-replicate survival statistics and DEG tables are in supplementary tables. Strains are available through CGC or on request.
Limitations (4)
  • The timing, magnitude or duration of hsp-16.2 induction may not faithfully mimic endogenous TDP regulation during cryptobiosis. · stated by the authors
  • Skeletonization-based mitochondrial morphology measurements may not resolve complex networks under extensive fusion or fragmentation. · stated by the authors
  • Hyperosmotic NaCl exposure with manually counted polyQ foci serves as a proxy for desiccation. True desiccation tolerance was not tested, and representative images were chosen for figures. · noted in this summary
  • Expression level or copy number of the transgenes is not compared across the four TDP lines in the text, so differences between proteins could partly reflect dosage. · noted in this summary

Read this if you are considering heterologous expression of tardigrade stress proteins for organismal stress tolerance or longevity, and want whole-animal somatic C. elegans data.

Citations
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Abstract

Tardigrades survive extreme environmental stress by expressing intrinsically disordered proteins that stabilize cellular structures during cryptobiosis. Because these proteins protect cells from desiccation, oxidative stress, and DNA damage, they have attracted interest as potential therapeutics to enhance stress resilience in other organisms. However, most studies have relied on cultured cells or germline expression, leaving their effects in somatic tissues largely unexplored. Here, we generated Caenorhabditis elegans strains expressing four tardigrade intrinsically disordered proteins, Damage Suppressor (DSUP), cytoplasmic-abundant heat-soluble (CAHS), mitochondrial-abundant heat-soluble (MAHS), and Ramazzottius varieornatus late embryogenesis abundant (RvLEAM), throughout somatic tissues. Surprisingly, constitutive expression of each protein reduced lifespan, impaired motility and reproduction, and failed to improve resistance to most environmental stressors. DSUP toxicity was tissue- and developmental stage-dependent, indicating that both spatial and temporal expression influence its physiological effects. Despite these detrimental phenotypes, several proteins reduced protein aggregation during desiccation-associated stress, demonstrating context-dependent protective effects. Together, our findings reveal that tardigrade stress proteins impose physiological tradeoffs in vivo and underscore the importance of evaluating candidate stress-protective proteins in physiologically relevant somatic models before therapeutic translation.

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