Homologue T24F1.2, which we named SAMP-1. The mammalian putative orthologue was originally identified within a proteomic screen for integral components on the inner nuclear membrane and named NET5 (Schirmer et al., 2003). NET5 was subsequently named Samp1 and shown to play a role in positioning nuclei in polarizing NIH 3T3 cells. Nuclear migration in polarizing mouse NIH3T3 cells relies on SUN-KASH bridges to couple moving actin arrays inside the cytoplasm for the nucleoskeleton (Luxton et al., 2010; Folker et al., 2011). This nuclear migration also demands Samp1, which partially colocalizes and coimmunoprecipitates with SUN proteins in transmembrane actin-associated nuclear (TAN) lines (Borrego-Pinto et al., 2012). The homologous protein in Schizosaccharomyces pombe, Ima1, interacts in yeast two-hybrid assays with all the SUN protein Sad1 and has been implicated within the maintenance of nuclear morphology (Hiraoka et al., 2011). Previously a broad bioinformatics study predicted that C. elegans SAMP-1 could be a component of the nuclear Calcitriol Impurities A site envelope and confirmed this localization in the early embryo applying a transgenic SAMP-1::GFP fusion protein (Gunsalus et al., 2005). However, nothing else is recognized concerning the function of C. elegans SAMP-1. WeSUN amin interactions to move nucleiFIGURE six: samp-1(RNAi) animals have a weak nuclear migration defect. (A ) Embryos have been stained for SAMP-1 localization. Lateral views, with anterior left and dorsal up. Scale bars, 10 m. For each pair of photos, SAMP-1 immunostaining is shown in white on the left and in red on the suitable when it really is merged with DAPI staining of nuclei in blue. (A, B) An early wild-type embryo. (C, D) A later, pre omma-stage embryo. Arrowhead points to PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/21269259 a hyp7 precursor nucleus. (E, F) A samp-1(tm2710)-null embryo is shown to demonstrate specificity in the antibody. (G) Numbers of nuclei inside the dorsal cords of wild-type or samp-1(tm2710)(+); samp-1(RNAi) L1 larvae. Every single gray dot represents an individual animal. The imply and 95 CI error bars are shown. (H, I) DIC and GFP images showing two hyp7 nuclei abnormally within the dorsal cord (arrows) of a samp-1(tm2710)(+); samp-1(RNAi) L1 larva. The dorsal cord is up and is demarcated by the dotted line. Scale bar, 10 m.hence set out to examine the function of C. elegans SAMP-1 in nuclear migration. We first characterized the intracellular localization pattern of endogenous SAMP-1 to find out whether or not it was plausible that SAMP-1 functions at the nuclear envelope through nuclear migration in embryonic hyp7 precursor. Antibodies were raised against the C-terminus of 
SAMP-1. Anti AMP-1 antibodies recognized a band on the predicted size on a Western blot. The band intensity was tremendously lowered in samp-1(RNAi) extracts (Supplemental Figure S2). SAMP-1 antibodies localized strongly to a ring about 4,6-diamidino-2-phenylindole (DAPI) tained nuclei, constant with nuclear envelope staining, in all cells of wild-type early embryos but not in samp1(tm2710) likely null embryos (Figure six and Supplemental Figure S2). Thus the antibody is specific for SAMP-1, having a localization pattern anticipated for a nuclear membrane protein. While we did not test the certain localization within the nuclear envelope, we hypothesize that SAMP-1 is definitely an inner nuclear membrane protein determined by the published localization on the mouse orthologue, Samp1 (Buch et al., 2009). In later embryos in the time of hyp7 nuclear migration, SAMP-1 localization at the nuclear envelope was less strong and restricted.
