Extended Data Fig. 3: Transgenerational regulation of GLD-1 expression by mstr genes.
From: Noncanonical inheritance of phenotypic information by protein amyloids
(a) STRING interaction network of proteins detected in proximity to MSTR-1 by BioID. Red circle, highly connected interactors by heat diffusion. Green highlights, 19S proteasomal regulatory subunits; orange highlight, 20S core proteasomal subunit. (b) Schematic of the mstr-1 locus and alignment of the Alphafold structures of C. elegans MSTR-1 and human ZFAND5. Plot, brood size of the mstr-1(ok1685) allele which only ablates the A20 domain. Lines, mean (± S.D.). ns, not significant for genotype effect. Two-way ANOVA with Bonferroni’s correction. n = 31 mstr-1(ok1685) and 26 mstr-1(ok1685); mstr-2 worms. (c) Western blot of immunoprecipitated MSTR-1::FLAG. FK2, antibody used to detect mono- and poly-ubiquitin conjugated proteins. M2, FLAG antibody. Red arrowhead, band corresponding to MSTR-1. (d) Representative CLIP-seq experiment showing regions excised for library preparation and sequencing. (e) Left, MA plot of CLIP-seq counts of reads mapping to each transcript. Blue dots, significantly enriched in respective sample (P < 0.02, Benjamini–Hochberg-adjusted Wald test). Red dots, significantly enriched germ line sex determining genes (SDG) deduced from a GO enrichment analysis (GO: 0018992, FE = 6.22, Fisher’s exact P = 0.000811, Benjamini–Hochberg FDR = 0.0157). Right, comparison of statistically significant (P < 0.02) transcripts enriched in MSTR-1 CLIP-seq (X-axis) to transcripts significantly differentially expressed in mstr-1 ablated worms. n = 2 MSTR-1 pulldowns and 1 control pulldown. (f) Immunoprecipitation of GLD-1::V5 followed by Western blot using anti-ubiquitin (left) and V5 (right) antibodies. (g) GLD-1 expression in wild-type and mstr worms at late generation 20 °C. Bar, 20 μm. (h, i) Transferring worms to 25 °C leads to increased penetration of ectopic GLD-1::mScarlet expression over a small number of generations. C, P0 at 25 °C, D, F3 at 25 °C. Transgenerational ectopic expression of GLD-1 in mstr worms is rescued by the rpt-1 suppressor. Exact n values are provided in Source Data. Bar, 20 μm. Uncropped blots, numerical data and statistical values are available in source data.
