Cloning of a rat cDNA encoding a novel LIM domain protein with high homology to rat RIL

A complementary DNA (1392 bp) encoding a protein with high homology to rat reversion-induced LIM (RIL) protein was cloned from rat hepatocytes by differential screening of a subtractive (normoxic minus hypoxic) lambda GEM-2 cDNA library. This cDNA clone, denoted CLP-36, encodes a 327-amino-acid (aa) protein that contained a restrictively conserved LIM (a Cys-rich domain with consensus aa sequence C-X2-C-X17-19-H-X2-C-X2-C-X2-C-X16-20-C-X2-C/H/D that was initially identified in homeodomain proteins, Lin-11 [Freyd et al., Nature 344 (1990) 876-879], Isl-1 [Karlsson et al., Nature 344 (1990) 879-882] and Mec-3 [Way et al., Cell 54 (1988) 5-16] in the C-terminal portion (aa 192-327). It shared an overall 45.1% identity to a rat LIM domain RIL protein [Kiess et al., Oncogene 10 (1995) 61-68], with 62.0% identity in the N terminus (aa 1-89) and 50.0% identity in the C terminus (aa 188-327). The rat CLP-36 mRNA is expressed most abundantly in heart, lung and liver, moderately in spleen and skeletal muscle, and at extremely low levels (if at all) in testis and brain tissues. Northern blotting analysis of total RNA extracted from normoxic or hypoxic rat hepatocytes, with a fragment of clone CLP-36 as probe, demonstrated a single band with a mobility corresponding to a size of 1.4 kb, whose level was significantly decreased during chemical hypoxia.