Intracellular hyaluronic acid-binding protein 4 (HABP4): a candidate tumor suppressor in colorectal cancer.

Talita Diniz Melo-Hanchuk,Jörg Kobarg,Ângela Saito,Carolina Colleti,José Barreto Campello Carvalheira,Jose Vassallo,Maria Carolina Santos Mendes

doi:10.18632/oncotarget.27804

Abstract

Hyaluronic Acid-binding protein 4 (HABP4) is a regulatory protein of 57 kDa that is functionally involved in transcription regulation and RNA metabolism and shows several characteristics common to oncoproteins or tumor suppressors, including altered expression in cancer tissues, nucleus/cytoplasm shuttling, intrinsic lack of protein structure, complex interactomes and post translational modifications. Its gene has been found in a region on chromosome 9q22.3-31, which contains SNP haplotypes occurring in individuals with a high risk for familial colon cancer. To test a possible role of HABP4 in tumorigenesis we generated knockout mice by the CRISPR/Cas9 method and treated the animals with azoxymethane (AOM)/dextran sodium sulfate (DSS) for induction of colon tumors. HABP4–/– mice, compared to wild type mice, had more and larger tumors, and expressed more of the proliferation marker proteins Cyclin-D1, CDK4 and PCNA. Furthermore, the cells of the bottom of the colon crypts in the HABP4–/– mice divided more rapidly. Next, we generated also HABP4–/– HCT 116 cells, in cell culture and found again an increased proliferation in clonogenic assays in comparison to wild-type cells. Our study of the protein expression levels of HABP4 in human colon cancer samples, through immunohistochemistry assays, showed, that 30% of the tumors analyzed had low expression of HABP4. Our data suggest that HABP4 is involved in proliferation regulation of colon cells in vitro and in vivo and that it is a promising new candidate for a tumor suppressor protein that can be explored both in the diagnosis and possibly therapy of colon cancer.

Highlights

Colon cancer is an important contributor to worldwide cancer morbidity and mortality
Our data suggest that Hyaluronic Acid-binding protein 4 (HABP4) is involved in proliferation regulation of colon cells in vitro and in vivo and that it is a promising new candidate for a tumor suppressor protein that can be explored both in the diagnosis and possibly therapy of colon cancer
To knockout mouse HABP4 gene through the CRISPR/Cas9 technique, a 20-nt single guide RNA (sgRNA) targeting the first exon of Habp4 was designed, cloned into the px330 vector, which was subsequently injected into C57BL/6J mouse zygotes

Summary

Introduction

Colon cancer is an important contributor to worldwide cancer morbidity and mortality. In an analysis of familial colon cancer risk-associated SNP haplotypes, the human gene encoding the protein www.oncotarget.com intracellular hyaluronic acid-binding protein 4 (HABP4) [4], has been found in linkage disequilibrium [5]. The Ki-1 antibody recognizes aside the surface CD30 transmembrane protein a 57 kDa cytoplasmic and nuclear antigen [8, 9]. The full length cDNA cloning revealed that this 57 kDa protein, rich in positively charged amino acids in its sequence, interacts in vitro with negatively charged macromolecules such as RNAs and hyaluronic acid, among others, resulting in its denomination as Hyaluronic acid binding protein 4 (=HABP4) [4]. Several functional studies performed later, pointed to HABP4 as intracellular regulatory protein, that is rather involved in expression regulation on various levels [10,11,12,13,14,15]

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Conclusion