Zebrafish P54 RNA helicases are cytoplasmic granule residents that are required for development and stress resilience

Abstract

Stress granules are cytoplasmic foci that directly respond to the protein synthesis status of the cell. Various environmental insults, such as oxidative stress or extreme heat, block protein synthesis; consequently, mRNA will stall in translation, and stress granules will immediately form and become enriched with mRNAs. P54 DEAD box RNA helicases are components of RNA granules such as P-bodies and stress granules. We studied the expression, in cytoplasmic foci, of both zebrafish P54 RNA helicases (P54a and P54b) during development and found that they are expressed in cytoplasmic granules under both normal conditions and stress conditions. In zebrafish embryos exposed to heat shock, some proportion of P54a and P54b helicases move to larger granules that exhibit the properties of genuine stress granules. Knockdown of P54a and/or P54b in zebrafish embryos produces developmental abnormalities restricted to the posterior trunk; further, these embryos do not form stress granules, and their survival upon exposure to heat-shock conditions is compromised. Our observations fit the model that cells lacking stress granules have no resilience or ability to recover once the stress has ended, indicating that stress granules play an essential role in the way organisms adapt to a changing environment.

rbpms2 functions in Balbiani body architecture and ovary fate

“…Wild-type GFP-Rbpms2 was detected near the nucleus (~20%) and associated with the centrosomes or spindle (9%), and as in HEK293 cells, GFP-Rbpms2 was localized to granules in 65% of somatic cells of the zebrafish embryo (Figs 3G–3H and 4 1; n = 489 cells, 19 embryos). To determine the identity of these Rbpms2 positive granules, we examined endogenous markers, including the classical stress granule marker Tial-1 and the p-body marker Dcp2, using antibodies previously validated in zebrafish [ 29 ]. We detected no overlap with Tial-1 ( Fig 3G ; n = 172 cells, 8 embryos) and partial overlap with Dcp2 ( Fig 3H ; n = 317 cells, 11 embryos), indicating that the granules to which GFP-Rbpms2 localizes in zebrafish somatic cells are not stress granules, but that a small subset are GFP-Rbpms2 and Dcp2 positive granules.…”

“…Rabbit anti-ACF7/Macf1 was used at 1:1000 [ 52 ]. Rabbit anti-DCP2 (Novus Biologicals, NBP2-16109) and rabbit anti- TIAL-1 (Novus Biologicals, NBP1-79932) were used at1:2000 as in [ 29 ]. Alexafluor488, Alexafluor568, CY3, C5 (Molecular Probes) secondary antibodies were diluted at 1:500.…”

Optogenetic modulation of TDP-43 oligomerization accelerates ALS-related pathologies in the spinal motor neurons

“…We also tested whether components of conventional stress granules (SGs) were recruited to these aggregates by using the antibodies against human G3BP and TIAL1 that robustly co-labeled heat shockinduced SGs in zebrafish ( Supplementary Fig. 6) 39,40 . The immunoreactivity for anti-G3BP antibody was detected in about 65% of the cells containing large aggregates of opTDP-43h or opTDP-43h A315T (Fig.…”

YB-1 recruitment to stress granules in zebrafish cells reveals a differential adaptive response to stress

“…The zebrafish ( Danio rerio ) is a fresh-water fish of the cyprinid family, and it represents one of the most popular and versatile genetic models for environmental and human disease studies 21,22 . However, in zebrafish, only limited information concerning the presence and regulation of SG-like structures is available 23–25 . We have recently shown that the subcellular localization of YB-1 is regulated by the circadian clock in zebrafish 26 .…”