Genomic Structure of Human OKL38 Gene and Its Differential Expression in Kidney Carcinogenesis

Abstract

We previously demonstrated the growth inhibitory property of OKL38 and its possible roles in mammary carcinogenesis. To further understand the regulation and roles of OKL38 in tumorigenesis we proceeded to clone and characterize the human OKL38 gene and three of its variants with transcripts of 1.9, 2.2, and 2.4 kb. The human OKL38 gene spans ϳ18 kb and contains 8 exons and 7 introns with exon size ranging from 92 to 1270 bp. RT-PCR and sequence analysis suggest that different transcripts were arrived through differential promoter usage and alternate splicing. Multiple Tissue Expression array (MTE) and Multiple Tissue Northern blot (MTN) indicated that OKL38 was ubiquitously expressed in all tissues with high expression in liver, kidney, and testis. The cancer profiling array (CPA) of paired normal/tumor cDNA showed that OKL38 mRNA was down-regulated in 70% (14 of 20) of kidney tumors. Western analysis revealed that the OKL38 protein was undetectable in 78% (7 of 9 pairs) of kidney tumor tissues. Immunohistological analysis showed that 64% (14 of 22) of kidney tumors were either lost or underexpressed OKL38 protein compared with the adjacent normal tissue. A transfection study using OKL38-eGFP recombinant construct showed that overexpression of the 52 kDa OKL38 protein in A498 cells resulted in growth inhibition and cell death. This study demonstrates the complex genomic structure of the OKL38 gene and its growth inhibitory and cytotoxic properties. Our data suggest the potential use of OKL38 in diagnosis, prognosis, and/or treatment of kidney cancer.

The role of 5′ untranslated region in translational suppression of OKL38 mRNA in hepatocellular carcinoma

“…Conversely, Riou et al (2002) have reported the overexpression of OKL38 transcripts in HCC, even though the gene resided in chromosome 16q24, a region prone to loss of heterozygosity. Re-establishing the expression of HuOKL38-1a protein was shown to cause rounding and subsequently cell death of Chang liver cells, which was similar to our previous studies in kidney cancer (Ong et al, 2004a). Importantly, we have demonstrated that the 38 kDa isoform was more potent in inducing cell death compared to the 52 kDa isoform (Figure 3a).…”

“…This observation was in agreement with our previous study in A498 kidney cancer cells (Ong et al, 2004a). Based on the above observations, we postulated that the discrepancy between mRNA and protein levels could be attributed to (i) mutation within the coding region leading to frame shift or change of amino-acid sequence, which resulted in the loss of protein expression or protein produced but not detected by anti-OKL38 antibody; and (ii) the involvement of the UTRs in translational regulation of OKL38 mRNA.…”

“…The loss of OKL38 protein (38 kDa isoform) is significantly correlated with the tumor stages of the disease, which is in agreement with low level of OKL38 transcripts observed in breast cancer cell lines (Huynh et al, 2001) and the loss of OKL38 protein in kidney cancer (Ong et al, 2004a). Conversely, Riou et al (2002) have reported the overexpression of OKL38 transcripts in HCC, even though the gene resided in chromosome 16q24, a region prone to loss of heterozygosity.…”

“…Four OKL38 variants have previously been identified, of which three were cloned (Ong et al, 2004a). The same ORF that encoded for a 52 kDa protein was found in HuOKL38-1a and -2a variants, whereas the HuOKL38-2b variant translates a larger protein of 61 kDa.…”

“…OKL38 mRNA is significantly reduced in breast cancer cell lines, 7,12-dimethylbenz[a]anthracene-induced breast cancer (Huynh et al, 2001) and kidney tumors (Ong et al, 2004a). On the contrary, OKL38 transcripts were reported to be overexpressed in HCC and adjacent benign tissue as compared to healthy normal liver tissue (Riou et al, 2002).…”