Hepatoma-derived Growth Factor Stimulates Cell Growth after Translocation to the Nucleus by Nuclear Localization Signals

Abstract

Hepatoma-derived growth factor (HDGF) is the original member of the HDGF family of proteins, which contains a well-conserved N-terminal amino acid sequence (homologous to the amino terminus of HDGF; hath) and nuclear localization signals (NLSs) in gene-specific regions other than the hath region. In addition to a bipartite NLS in a gene-specific region, an NLS-like sequence is also found in the hath region. In cells expressing green fluorescence protein (GFP)-HDGF, green fluorescence was observed in the nucleus, whereas it was detected in the cytoplasm of cells expressing GFP-HDGF with both NLSs mutated or deleted. GFP-hath protein (GFP-HATH) was distributed mainly in the nucleus, although some was present in the cytoplasm, whereas GFP-HDGF with a deleted hath region (HDGFnonHATH) was found only in the nucleus. Exogenously supplied GFP-HDGF was internalized and translocated to the nucleus. GFP-HATH was internalized, whereas GFP-HDGFnonHATH was not. Overexpression of HDGF stimulated DNA synthesis and cellular proliferation, although HDGF with both NLSs deleted did not. Overexpression of HDGFnonHATH caused a significant stimulation of DNA synthesis, whereas that of hath protein did not. HDGF containing the NLS sequence of p53 instead of the bipartite NLS did not stimulate DNA synthesis, and truncated forms without the C- or N-terminal side of NLS2 did not. These findings suggest that the gene-specific region, at least the bipartite NLS sequence and the N- and C-terminal neighboring portions, is essential for the mitogenic activity of HDGF after nuclear translocation.

Role of hepatoma‐derived growth factor in promoting de novo lipogenesis and tumorigenesis in hepatocellular carcinoma

“…The consensus sequence of the bidirectional nuclear localization signal is found in both the PWWP domain and other regions of the HDGF sequence. Nuclear localization of HDGF is essential for the mitogenic activity of HDGF in cells(Kishima et al ., 2002). Our results suggest that the location of HDGF is strongly related to the poor prognosis of patients with HCC.…”

Hepatoma-derived Growth Factor

“…5C). These results clearly demonstrate that HDGF harbors two different proliferative activities: one localized inside the carboxyl-terminal non-HATH region responsible for intranuclear activity (26,37), and the other located in the HATH81-100 region exerting its mitogenic activity from the cell surface.…”

“…Recent studies on HEK 293 cells demonstrated that a GFP-HDGF fusion protein added to the cell culture medium can be internalized and is able to translocate into the nucleus within 72 h after addition (26). We added a HDGF-GFP and HDGF-␤-galactosidase fusion protein to the cell culture medium of primary human fibroblasts and NIH 3T3 cells for time periods of up to 72 h. Cells were investigated by fluorescence microscopy or ␤-galactosidase activity determinations.…”

“…HDGF has two nuclear localization signals, one in the conserved HATH region, and the other in the carboxyl-terminal area specific for the different family members. The nuclear localization has been shown to be a prerequisite for the mitogenic activity of intracellularly expressed HDGF (5,26). Several groups have shown independently that HDGF is mitogenic not only when expressed in cells but also when applied exogenously (2,3).…”

“…Current data for these FGF family members suggest that these growth factors may be released from cells via a mechanism independent of the secretory endoplasmic reticulum-Golgi-plasma membrane pathway (27)(28)(29)(30). For exogenously applied HDGF and its related protein, LEDGF, it has also been shown that they can be internalized, and it has been proposed that the mitogenic action of exogenous HDGF depends on internalization and subsequent nuclear translocation (26,31). Internalization would require a cell surface receptor, and in this respect, it is important to note that HDGF strongly binds heparin-like glycosaminoglycans (HLGAGs) (4,10).…”