The high mortality in breast cancer is often associated with metastatic progression in patients. Previously we have demonstrated that testes-specific protease 50 (TSP50), an oncogene overexpressed in breast cancer samples, could promote cell proliferation and tumorigenesis. However, whether TSP50 also has a key role in cell invasion and cancer metastasis, and the mechanism underlying the process are still unclear. Here we found that TSP50 overexpression greatly promoted cell migration, invasion, adhesion and formation of the stellate structures in 3D culture system in vitro as well as lung metastasis in vivo. Conversely, TSP50 knockdown caused the opposite changes. Mechanistic studies revealed that NF-κB signaling pathway was required for TSP50-induced cell migration and metastasis, and further results indicated that TSP50 overexpression enhanced expression and secretion of MMP9, a target gene of NF-κB signaling. In addition, knockdown of MMP9 resulted in inhibition of cell migration and invasion in vitro and lung metastasis in vivo. Most importantly, immunohistochemical staining of human breast cancer samples strongly showed that the coexpression of TSP50 and p65 as well as TSP50 and MMP9 were correlated with increased metastasis and poor survival. Furthermore, we found that some breast cancer diagnosis-associated features such as tumor size, tumor grade, estrogen receptors (ER) and progesterone receptors (PR) levels, were correlated well with TSP50/p65 and TSP50/MMP9 expression status. Taken together, this work identified the TSP50 activation of MMP9 as a novel signaling mechanism underlying human breast cancer invasion and metastasis.
ABSTRACT. We investigated the effects of pulsed electromagnetic fields (PEMFs) of 20 Hz/2 mT on the osteogenic and adipogenic differentiation of bone marrow stem cells (BMSCs). Sprague Dawley rat BMSCs were isolated and cultured in vitro. The BMSCs of the third passage were obtained and stimulated by PEMFs of 20 Hz/2 mT. The alkaline phosphatase (ALP) activity was measured according to the ALP assay kit manufacturer instructions, the BMSC osteogenic and adipogenic indicators were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR), and oil red O staining was used to observe the adipose-induced adipogenic differentiation of BMSCs. PEMFs of 20 Hz/2 mT significantly promoted the activity of ALP in the BMSCs (P < 0.01) and mRNA expression of osteogenic proteins (osteocalcin and osteopontin). The PEMFs inhibited the expression of adipogenic transcription factors such as adipokines and T. Lu et al. 11536©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 14 (3): 11535-11542 (2015) adipocyte-binding protein-2, and the adipogenic differentiation of BMSCs. PEMFs of 20 Hz/2 mT can promote osteogenic differentiation and inhibit adipogenic differentiation in BMSCs.
A 1446 bp SalI fragment of LsNPV was sequenced by the silver staining method, and two large open reading frames (ORFs, ORF1 and ORF2) were found, both contain typical characteristics of the 5' regulatory elements of baculovirus early genes. ORF1 is 345 bp long with the capacity to encode a putative protein of 114 amino acid residues with MW about 13 kDa and was designated p13 gene, ORF2 comprises 248 bp from the 3' end of the fragment. In the untranslated region (UTR) of ORF1, a 33 bp mini cistron (ORF3), a core recognition sequence (CGTCG) for many bHLHzip transcription factors and a late promoter sequence TTAAG are present. In the UTR of ORF2, two host transcription factor binding elements (CACGTG and GATA motif) and two CGT motifs were found. Some regular leucine zipper-like structures, designated leucine trans-conformation structure and LVT repeat, were found near the N-terminus and the middle of p13 protein. The leucine trans-conformation structure that is near the N-terminus consists of 4 leucines and 7 other amino acids between every two leucines, and every leucine is located at a conformation shift point of the predicted secondary structure of the p13 protein. In LVT repeat, L-6aa-V-6aa-T-6aa is repeated once. The functions of those structures remain unclear, and the two ORFs, not found in the genome of Autographa californica nuclear polyhedrosis virus, are possibly two new genes.
Testes-specific protease 50 (TSP50), a novelly identified oncogene, has the capacity to induce cell proliferation, cell invasion and tumor growth. Further studies indicated that CAGA-luc (an activin-responsive reporter construct) reporter activity could be significantly suppressed by TSP50 overexpression, implying that the activin signaling may participate in TSP50-mediated cell proliferation. Here, we reported that TSP50 had an inhibitory effect on activin signaling. Mechanistic studies revealed that TSP50 could interact with ActRIIA, inhibit activin typeIreceptor (ActRIB) phosphorylation, repress Smad2/3 nuclear accumulation and finally promote cell proliferation by reducing the expression of activin signal target gene p27. Additionally, we found that ActRIB activation could reverse TSP50-mediated cell proliferation and tumor growth. Furthermore, analysis of human breast cancer specimens by immunohistochemistry indicated that TSP50 expression was negatively related to p-Smad2/3 and p27 protein levels. Most importantly, breast cancer diagnosis-related indicators such as tumor size, tumor grade, estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor 2 (HER-2) levels, were correlated well with TSP50/p-Samd2/3 and TSP50/p27 expression status. Thus, our studies revealed a novel regulatory mechanism underlying TSP50-induced cell proliferation and provided a new favorable intervention target for the treatment of breast cancer.
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