Overexpression of humbug promotes malignant progression in human gastric cancer cells

Abstract

Abstract. Two human gastric cancer cell lines of differing invasive potential, SNU-484 and SNU-638 cells, were examined using subtractive suppression hybridization in a search for any genes associated with metastasis. Of the eight cDNAs identified as being differentially expressed genes, it was determined that humbug, which encodes a truncated isoform of aspartyl (asparaginyl) ß-hydroxylase (AAH) missing catalytic domain, was overexpressed in highly invasive SNU-638 cells. Expression analysis showed that the mRNA expression level of humbug was correlated with invasive potential in various human gastric cancer cell lines. The forced expression of humbug to the human gastric cancer cell line AGS increased its anchorage-independent growth in 0.3% agar without affecting cell proliferation. Furthermore, humbug-transfected cells migrated more actively and showed an increased invasion rate relative to vector-transfectants or parental AGS in vitro. This is the first demonstration that humbug, a truncated form of AAH, can be overexpressed during the malignant progression of human gastric cancer cells and that it can function as a metastasis-inducing gene.

Prognostic value of aspartyl (asparaginyl)-β-hydroxylase/humbug expression in non–small cell lung carcinoma

“…Humbug’s probable role in regulating intracellular calcium through ryanodine receptors in the endoplasmic reticulum [15–16] is likely critical to the assembly and disassembly of cellular cytoskeletal proteins and, consequently, the modulation of cell shape and adhesion. Indeed, recent in vitro studies showed humbug-transfected gastric cancer cells to migrate more robustly than the nontransfected counterparts [17]. Thus, overexpression of ABH and humbug may have a synergistic effect on the dynamic alterations in cell shape and adhesion, which are essential to the processes of invasion and metastatic spread of tumors.…”

“…ABH catalyzes hydroxylation of specific aspartyl and asparaginyl residues in epidermal growth factor–like domains of proteins such as Notch, Jagged, and cellular adhesion molecules such as laminin and tenascin [9–12]. Humbug is a truncated homolog of ABH [13,14] and lacks ABH’s catalytic domain that has roles in intracellular calcium regulation [15,16] and cell motility [17], which is required for metastatic spread of malignant neoplasms.…”

Potential Role of Phosphorylation as a Regulator of Aspartyl-(asparaginyl)-β-hydroxylase: Relevance to Infiltrative Spread of Human Hepatocellular Carcinoma

“…AAH is physiologically cleaved into a ∼30-34 kD N-terminal fragment that is identical to Humbug, a truncated isoform that binds to calcium and promotes adhesion [21,22], and a ∼52-56 kD catalytically active C-terminal domain that contains a 675- His residue that is essential for hydroxylation activity [23]. AAH hydroxylates β-carbons of specific aspartate (Asp) and asparagine (Asn) residues within consensus sequences of epidermal growth factor (EGF)-like domains of targets [24] such as Notch and Jagged [11,25,26].…”

Development of a novel anti-human aspartyl-(asparaginyl) β-hydroxylase monoclonal antibody with diagnostic and therapeutic potential

“…As such, the development of an efficient detection method is critical. Several hosts, including E. coli and various mammalian cell lines, have been used to express the recombinant subtype proteins of HAAH (2,17). A previous investigation also used the pProEX expression system to express humbug (a subtype of HAAH) in E. coli (11).…”