TY  - GEN
T1  - Visualization of endogenous NID-1 and EMB-9 in C. elegans
AU  - Matsuo, Kanae
AU  - Koga, Akihiro
AU  - Ihara, Shinji
DO  - 10.17912/micropub.biology.000110
UR  - http://beta.micropublication.org/journals/biology/micropub-biology-000110/
AB  - nid-1 and emb-9 are the worm homologs of the human nidogen and human type IV collagen, respectively (Kramer, J. M. 2005). Nidogen is a glycoprotein that can bind type IV collagen with high affinity. nid-1 is single nidogen gene in the C. elegans genome, generating three alternative splice variants (nid-1A, nid-1B, and nid-1C) (Kang & Kramer, 2000). Type IV collagen is the principal component of basement membrane (BM), a complex network of triple helical molecules. emb-9 encodes sole α1-like type IV collagen that forms a heterotrimer with one α2-like type IV collagen (let-2). Nidogen and type IV collagen are components of BM that cover the basal surfaces of nearly all animal tissues. Previous work in C. elegans has confirmed the localizations of NID-1 and EMB-9 by using a transgenic strain or immunohistochemistry (Graham et al. 1997; Kim & Wadsworth, 2000) . To address endogenous localization patterns of both proteins in vivo, we used CRISPR/CAS9 technology to insert fluorescence label at the nid-1 and emb-9 genomic locus, and established worms expressing the mkate2::NID-1 or EMB-9::mcherry, respectively. To visualize all isoforms, nid-1A, B, and C, we inserted mkate2 into exon 2, which is a common exon for all isoforms. We found that mkate2::NID-1strongly localized to intestinal BM (Figure 1B and E) and localized to gonadal BM and nerve ring (Figure 1E and G), consistent with previous reports. Interestingly, we found that EMB-9::mcherry strongly localized to BM in pharynx (Figure 1D and F), although mkate2::NID-1 was faintly localized at pharyngeal BM (Figure 1E). We also confirmed strong expression of EMB-9::mcherry in coelomocytes, but not in the strain expressing mkate2::NID-1 (Figure 1G and H). The mkate2::NID-1 and EMB-9::mcherry are similarly localized to BM at gonad and uterus (Figure 1G and 1H). Differences of localization intensity at BM in the tissues are summarized in Figure 1G. Our work indicated that endogenous NID-1 and EMB-9 show different intensities at BM of pharynx, intestine, although it remains unclear whether these differences are associated with organ function.
PY  - 2019
JO  - microPublication Biology
ER  -