Nucleoplasmic calcium regulates cell proliferation through legumain

Abstract

Background and Aims Nucleoplasmic Ca2+ regulates cell growth in the liver, but the proteins through which this occurs are unknown. Methods We used Rapid Subtraction Hybridization (RaSH) to subtract genes in SKHep1 liver cells expressing the Ca2+ buffer protein parvalbumin (PV) targeted to the nucleus, from genes in cells expressing a mutated form of nuclear-targeted PV which has one of two Ca2+-binding site inactivated. The subtraction permitted selection of genes whose expression was affected by a small alteration in nuclear Ca2+ concentration. Results The asparaginyl endopeptidase legumain (LGMN) was identified in this screening. When Ca2+ was buffered in the nucleus of SKHep1 cells, LGMN mRNA was decreased by 97%, in part by a transcriptional mechanism, and decreased expression at the protein level was observed by immunoblot and immunofluorescence. Treatment with Hepatocyte Growth Factor increased LGMN expression. Knockdown of LGMN by siRNA decreased proliferation of SKHep1 cells by ~50% as measured both by BrdU uptake and mitotic index, although an inhibitor of LGMN activity did not affect BrdU incorporation. A significant reduction in the fraction of cells in G2/M phase was seen as well. This was associated with increases in expression of cyclins A and E. Furthermore, LGMN expression was increased in hepatocellular carcinoma cells relative to normal hepatocytes in the same specimens. Conclusions These findings suggest a new role for LGMN and provide evidence that nuclear Ca2+ signals regulate cell proliferation in part through modulation of LGMN expression. Increased expression of LGMN may be involved in liver carcinogenesis.

Proteolysis mediated by cysteine cathepsins and legumain—recent advances and cell biological challenges

“…In comparison to the limited variety of biochemical cleavage mechanisms, there is an impressively diverse cellular and extracellular repertoire of physiological substrates, as well as endogenous protease inhibitors which are called into action for the regulation of proteolysis in response to cellular conditions (Andrade et al 2011;Brix et al 2013;Dauth et al 2011a;Turk et al 2008). Locally and temporally controlled protease activities are already seen very early in life, such as the acrosomal process during fertilization of the egg cell (Mao and Yang 2013).…”

Activation of Asparaginyl Endopeptidase Leads to Tau Hyperphosphorylation in Alzheimer Disease

“…Processing of AEP requires sequential removal of the C-and N-terminal propeptides at different pH thresholds to generate 46-kDa (active pro-AEP) and 36-kDa (active AEP) active enzymes (8,9). AEP has multiple roles depending on substrate specificity such as initiator of invariant chain processing during MHC class IImediated antigen presentation (10), inducer of cell migration (11,12), and modulator of processes such as proliferation (13) and both pro-death and pro-survival functions using apoptotic mechanisms (14). The variety of roles of AEP is due to its ability to proteolyse multiple substrates, including among others the extracellular matrix protein fibronectin (15), progelatinase (16), and cathepsin H, B, and L (17).…”

The expression of asparaginyl endopeptidase promotes growth potential in epithelial ovarian cancer

“…9,10 Moreover, some studies have reported that AEP was related to tumor cell proliferation. 8,23 And blocking of AEP with antibody was reported to inhibit tumor progression and metastasis. Once AEP combined with its substrate recognition, it could be upregulated in solid tumors.…”

“…Recently, it is reported that AEP is highly expressed in many solid tumors, such as hepatocellular cancer, gastric cancer, breast cancer, colorectal cancer and lung cancer. [8][9][10][11][12][13] However, the mechanism of AEP in epithelial ovarian cancer remains unknown.…”