Expression of prosalusin in human neuroblastoma cells

Abstract

Salusins, which are derived from the prosalusin precursor molecule, regulate hemodynamics, mitogenesis and atherogenesis. The preprosalusin gene is ubiquitously expressed, while the salusin-β peptide is present in systemic endocrine cells and the neuroendocrine system. However, the regulatory mechanisms for the preprosalusin gene and prosalusin expression remain to be investigated. Real-time quantitative RT-PCR and salusin-α radioimmunoassay revealed that the neuroblastoma cell line, SK-N-SH, exhibited marked upregulation of preprosalusin mRNA and salusin-α-like immunoreactivity (LI) when incubated under 2% serum condition. However, SK-N-SH cells released a limited amount of salusin-α-LI into the culture supernatant. Reverse-phase high performance liquid chromatography coupled with radioimmunoassay after extraction of proteins from the conditioned media using an octyl-silica column did not reveal a component that co-eluted with authentic salusin-α. Western blotting of the nuclear extracts from SK-N-SH showed the expression of prosalusin and its cleaved fragments, but not authentic salusin-α. Addition of Jak-2 inhibitors to growing SK-N-SH cells cultured under 10% serum condition resulted in increased salusin-α-LI expression. Suppression of Jak-2 mRNA using siRNAs upregulated intracellular salusin-α-LI, as detected by immunofluorescence. In summary, the preprosalusin gene and prosalusin protein are expressed in a neuroblastoma cell line and upregulated by reduced serum. The Jak-2 pathway may be involved in the regulation of salusin expression.