The UNC-83/UNC-84 LINC members are required for body wall muscle nuclei positioning in C. elegans

Abstract

From a mutagenesis screen in the nematode C. elegans we isolated the mutant bar18, showing an accumulation of muscle cell nuclei around the posterior pharyngeal bulb of the worm. Quantification of the overall amount of body wall muscle nuclei, based on the muscle-specific reporter myo-3p::gfp::NLS, revealed that the number of nuclei in bar18 mutants is unchanged compared to WT worms. The accumulation of muscle nuclei around the posterior pharyngeal bulb is due to a positioning defect, which can be precisely quantified by subdividing the worm into head, neck, and posterior body segments. Whole-genome sequencing revealed that bar18 animals carry a mutation in the KASHdomain gene unc-83 causing a premature STOP. An additional unc-83mutant allele recapitulates the phenotype, as does a mutant allele of UNC-84, a SUN-domain containing protein that interacts with UNC-83. UNC-83 and UNC-84 belong to a Linker of Nucleoskeleton and Cytoskeletonnuclear (LINC) complex that bridges the nuclear lamina with the cytoskeleton. SUN and KASH domain proteins are conserved in mammals and mutations in the corresponding genes have been linked to cancer, autism, muscular dystrophy and other diseases. Additionally, LINC complexes that function in nuclear migration have also been identified in mammals. We were able to rescue the unc-83 mutant phenotype by expressing the WT gene under a muscle-specific (myo-3p) promoter, demonstrating that the effect is cell autonomous. Mutations in either unc-83 or unc-84 have previously been linked to nuclear migration defects in P cells, intestinal cells and hyp7 hypodermal precursors but not in muscles. Whether the mis-positioning of muscle nuclei is due to migration or anchoring defects still needs to be determined. Introduction Thenuclear lamina is connected to the cytoskeleton via different ‘Linker of Nucleoskeleton and Cytoskeleton’ (LINC) complexes with a variety of functions. LINC complexes are widely conserved over various phyla, which include organisms such as plants, slime molds, yeast, roundworms, fruit flies and mammals. LINC complexes cross the nuclear membrane and are composed of SUN and KASH domain-containing proteins, which interact in the perinuclear space between the inner and outer nuclear membrane. KASH proteins are located at the outer nuclear membrane and may interact with actin filaments, microtubules (via dynein and kinesin), intermediate filaments (via spectrin), centrosomes and other cytoplasmic organelles. SUN proteins are located at the inner nuclear membrane and are associated with both chromatin and nuclear lamins. Functions include nuclear movement and anchoring, moving meiotic chromosomes and telomeres and sensing mechanic stimuli [1] [2] [3]. The KASH protein UNC-83 and the SUN protein UNC-84 form a LINC complex in C. elegans, which is required for migration of nuclei in P cells, intestinal cells and hyp7 hypodermal precursors, by recruiting dynein and kinesin-1 to the nuclear surface [4] [5] [6] [7] [8] [9]. Furthermore, UNC-84 has been implicated in maintaining the nuclear architecture of force-bearing cells, like body wall muscles [10]. Objective Our aim was to describe and quantify the novel observation of mis-positioned body wall muscle nuclei upon loss of the UNC-83/UNC-84 LINC complex and to address the question of whether the effect was cell autonomous or not. The UNC-83/UNC-84 LINC members are reqired for body wall muscle nuclei positioning in C. elegans DOI: 10.19185/matters.201805000009 Matters Select (ISSN: 2297-9239) | 2 a