thymosin-beta(4) and Prostatic-Neoplasms

In prostate cancer, the tumour microenvironment (TME) represents an important regulator of disease progression and response to treatment. In the TME, cancer-associated fibroblasts (CAFs) play a key role in tumour progression, however the mechanisms underpinning fibroblast-cancer cell interactions are incompletely resolved. Here, we address this by applying cell type-specific labelling with amino acid precursors (CTAP) and mass spectrometry (MS)-based (phospho)proteomics to prostate cancer for the first time.. Reciprocal interactions between PC3 prostate cancer cells co-cultured with WPMY-1 prostatic fibroblasts were characterised using CTAP-MS. Signalling network changes were determined using Metascape and Enrichr and visualised using Cytoscape. Thymosin β4 (TMSB4X) overexpression was achieved via retroviral transduction and assayed by ELISA. Cell motility was determined using Transwell and random cell migration assays and expression of CAF markers by indirect immunofluorescence.. WPMY-1 cells co-cultured with PC3s demonstrated a CAF-like phenotype, characterised by enhanced PDGFRB expression and alterations in signalling pathways regulating epithelial-mesenchymal transition, cytoskeletal organisation and cell polarisation. In contrast, co-cultured PC3 cells exhibited more modest network changes, with alterations in mTORC1 signalling and regulation of the actin cytoskeleton. The expression of the actin binding protein TMSB4X was significantly decreased in co-cultured WPMY-1 fibroblasts, and overexpression of TMSB4X in fibroblasts decreased migration of co-cultured PC3 cells, reduced fibroblast motility, and protected the fibroblasts from being educated to a CAF-like phenotype by prostate cancer cells.. This study highlights the potential of CTAP-MS to characterise intercellular communication within the prostate TME and identify regulators of cellular crosstalk such as TMSB4X.

Topics: Cell Communication; Cell Line, Tumor; Cell Movement; Fibroblasts; Humans; Male; Prostatic Neoplasms; Proteomics; Tumor Microenvironment

Thymosin beta4, a major G-actin-sequestering protein, is known to be involved in tumor metastasis. In the present study, we found that thymosin beta4 expression promotes the formation of actin-based pseudopodia-like extensions, associated with cell migration, in human prostate cancer LNCaP cells. Treatment with the phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin and Cdc42/Rac1/RhoA inhibitor Clostridium difficile toxin B significantly reduced pseudopodia formation in thymosin beta4-overexpressing LNCaP cells, suggesting that the pseudopodia formation by thymosin beta4 is probably involved in PI3K and Rho family pathway. We recently reported that thymosin beta4 expression is upregulated by androgen deprivation in prostate cancer cells. The increase in thymosin beta4 may be one of the causes of prostate cancer progression after androgen ablation therapy.

Topics: Androstadienes; Bacterial Proteins; Bacterial Toxins; Butadienes; cdc42 GTP-Binding Protein; Cell Line, Tumor; Cell Movement; Humans; Male; Microscopy, Phase-Contrast; Nitriles; Phosphoinositide-3 Kinase Inhibitors; Prostatic Neoplasms; Pseudopodia; rac1 GTP-Binding Protein; Reverse Transcriptase Polymerase Chain Reaction; rhoA GTP-Binding Protein; Sirolimus; Thymosin; Transfection; Wortmannin

A peptide with a sequence identical to rat thymosin beta(Tb)15 was reported to be upregulated in human prostate cancer. However, in this report we provide evidence that TbNB, initially identified in human neuroblastoma, is the only Tb isoform upregulated in human prostate cancer and that the Tb15 sequence is not present herein. In addition, we demonstrate that human TbNB has a higher affinity for actin in comparison to Tb4 and promotes cell migration. In combination, this experimentally validates TbNB as functional homologue of rat Tb15 in the human organism and clarifies the current composition of the human Tb family.

Topics: Actins; Amino Acid Sequence; Animals; Cell Line; Cell Movement; Humans; Male; Molecular Sequence Data; Prostatic Neoplasms; Rats; RNA, Messenger; Sequence Homology, Amino Acid; Thymosin; Up-Regulation

Topics: Actins; Animals; Antineoplastic Agents; Cell Physiological Phenomena; Cell Transformation, Neoplastic; Humans; Male; Melanoma, Experimental; Neoplasm Metastasis; Neovascularization, Pathologic; Prostatic Neoplasms; Thymosin; Up-Regulation

Zinc exhibits inhibitory effects on apoptosis, and a deficiency in this metal generally causes this type of cell death to occur. In the present study, we found that exposure to zinc results in necrosis of prostate carcinoma cells. When zinc acetate was added to LNCaP or PC-3 cells in monolayer culture, they began to detach from the culture dishes, and viability was lost after 4-8 h. Most of the cell death was found to be due to necrosis as determined by double staining with fluorescein-isothiocyanate-labeled annexin V and ethidium bromide, and by detection of hypodiploid cells. Associated with the induction of necrosis was an increase in low molecular-mass proteins, identified by HPLC analysis to be thymosin beta10, parathymosin and GAGE in LNCaP cells, and thymosin beta4, parathymosin and metallothionein in PC-3. The time course of the increase of thymosin beta10 in LNCaP cells and thymosin beta4 in PC-3 cells was consistent with that of appearance of cell detachment and dead cells. These results indicate that zinc can induce necrosis and suggest that production of proteins including beta-thymosins is involved in induction of processes leading to cell detachment.

Topics: Adenocarcinoma; Annexin A5; Apoptosis; Cell Adhesion; Cell Division; Cell Survival; Copper; DNA, Neoplasm; Humans; Male; Metallothionein; Necrosis; Neoplasm Proteins; Prostatic Neoplasms; Thymosin; Tumor Cells, Cultured; Zinc