Brigitte Lankat-Buttgereit

The tumour suppressor Pdcd4: recent advances in the elucidation of function and regulation

Pdcd4 (programmed cell death 4) has been known as a tumour suppressor gene and potential target for anticancer therapies for several years. Initially, Pdcd4 was identified as a gene that is up‐regulated during apoptosis, but its precise role still remains to be defined. However, there is increasing evidence that Pdcd4 levels influence transcription, as well as translation, modulate different signal transduction pathways and might act as a tumour suppressor. Interestingly, recent data suggest that Pdcd4 function may depend on cell type and/or genetic background. This review summarizes the current knowledge regarding the function and regulation of Pdcd4.

Localization of collagen mRNA in normal and scleroderma skin by in‐situ hybridization

Abstract. Scleroderma is a fibrotic disease occurring in a localized or systemic form. Disturbed regulation of connective tissue metabolism plays an important role in its pathogenesis. However, until now, most of the data available were obtained from studies of fibroblasts in culture and there is considerable doubt that fibroblasts in a monolayer reflect the in‐vivo situation. Using in‐situ hybridization with specific antisense RNAs on frozen sections of skin, cells were detected displaying enhanced messenger RNA levels for type I and type III collagen in patients with localized and systemic scleroderma. Activated fibroblastic cells were often located near blood vessels in the deep dermis of patients with early stages of the disease and were mostly surrounded by mononuclear cells. These findings are in agreement with the concept that the interaction of fibroblasts with ‘immunocompetent cells’ is crucial in the initial activation of connective tissue metabolism in fibrosis.

Co-localization of transforming growth factor beta 2 with alpha 1(I) procollagen mRNA in tissue sections of patients with systemic sclerosis.

The role of transforming growth factor beta 2 (TGF-beta 2) in the pathogenesis of systemic sclerosis (SSc) was investigated by in situ hybridization of skin biopsies from six patients with SSc. Two patients with acute systemic lupus erythematosis (SLE), one with acute dermatomyositis (DM), and three healthy individuals were used as controls. TGF-beta 2 mRNA was found to be co-localized with pro alpha 1(I) collagen expression around dermal blood vessels in all patients with the inflammatory stage of SSc, whereas there was no expression of either gene in the dermis of patients in the fibrotic stage, the SLE patients or the normal controls. These findings provide evidence that TGF-beta 2 released by inflammatory cells around blood vessels may play a role in mediating the collagen gene disregulation in fibrosis.

Programmed cell death protein 4 (pdcd4): A novel target for antineoplastic therapy?

Pdcd4 is a novel gene first identified as a differentially expressed protein during apoptosis. In the meantime not only the impact of Pdcd4 in programmed cell death but also an implication in transformation suppression by inhibition of protein translation is discussed. These features implicate a potential value of Pdcd4 as a molecular target in cancer therapy. This review summarizes the current knowledge about expression, structure and function of Pdcd4.

Localization of collagen α1(I) gene expression during wound healing by in situ hybridization

The cellular localization of collagen alpha 1(I) chain gene expression during wound healing in rats was investigated using in situ hybridization. Activation of collagen gene expression was first found within fibroblastic cells around the deep layers of the granulation tissue as early as 16 h post wounding. Heavily labeled cells were also detected near the intact wound edge and around hair follicles. At day 6 intense alpha 1(I) collagen gene expression was found within most cells of the granulation tissue and after day 8 most of the activity was localized to cells directly underneath the epidermis. 26 d after the induction of the wounds hardly any alpha 1(I) collagen gene expression could be demonstrated, which indicates a close, time-dependent control of collagen synthesis during repair processes.

Cloning and complete amino acid sequences of human and murine basement membrane protein BM-40 (SPARC, osteonectin)

Amino acid sequences of 285 and 286 residues, respectively, were deduced for mouse and human BM‐40 from cDNA clones isolated from expression libraries. The sequences showed 92% identity and were also essentially identical to those of bone osteonectin and of the parietal endoderm protein SPARC. About 60% of the mouse BM‐40 sequence was confirmed by Edman degradation. Two of the seven disulfide bonds were localized which apparently separate two distinct domains of mouse BM‐40.

Molecular cloning, functional expression, and signal transduction of the GIP-receptor cloned from a human insulinoma

Glucose‐dependent insulinotropic polypeptide (GIP) plays an important role in the regulation of postprandial insulin secretion and proinsulin gene expression of pancreatic β‐cells. This study demonstrates the molecular cloning of a cDNA for the GIP‐receptor from a human insulinoma λgt11 cDNA library. The cloned cDNA encoded a seven transmembrane domain protein of 466 amino acids which showed high homology (41%) to the human glucagon‐like peptide 1 (GLP‐1) receptor. Homology to the GIP receptor from rat or hamster was 79% and 81%, respectively. When transfected stably into fibroblast CHL‐cells a high affinity receptor was expressed which coupled to the adenylate cyclase with normal basal cAMP and increasing intracellular cAMP levels under stimulation with human GIP‐1–42 (EC50 = 1.29 × 10−13 M). The receptor accepted only human GIP 1–42 (K d = 1.93 ± 0.2 × 10−8 M) and porcine truncated GIP‐1–30 (K d = 1.13 ± 0.1 × 10−8 M) as high affinity ligands. At 1 μM, exendin‐4 and (9–39)amide weakly reduced GIP‐binding (25%) whereas secretin, glucagon, glucagon‐like peptide‐1, vasoactive intestinal polypeptide, peptide histidine‐isoleucine, and pituitary adenylyl cyclase activating peptide were without effect. In transfected CHL cells, GIP‐1–42 did not increase intracellular calcium. Northern analysis revealed one transcript of human GIP receptor mRNA with an apparent size of 5.5 kb. The exact understanding of GIP receptor regulation and signal transduction will aid in the understanding of the incretin hormones failure to exert its biological action at the pancreatic B‐cell in type II diabetes mellitus.

Pdcd4 inhibits growth of tumor cells by suppression of carbonic anhydrase type II

To identify new genes that are upregulated during apoptosis we previously cloned rat pdcd4. While the role of pdcd4 is still unclear it seems to possess a tumor suppressor activity. Pdcd4 directly interacts with the RNA helicase eIF4A and inhibits protein synthesis by interfering with the assembly of the cap-dependent translation initiation complex. In the present study, we show that pdcd4 suppresses carbonic anhydrase type II protein expression in HEK293 and Bon-1 carcinoid cells. Since tumor cells require a high bicarbonate flux for their growth, carbonic anhydrase suppression results in growth inhibition. Similar to pdcd4, carbonic anhydrase inhibitor ethoxyzolamide reduces growth of several endocrine tumor cell lines. Thus, the translation inhibitor pdcd4 represses endocrine tumor cell growth by suppression of carbonic anhydase II. Furthermore, carbonic anhydrase inhibitors might represent promising tools for anti-endocrine tumor treatment.

The transporter associated with antigen processing TAP: structure and function.

The transport of antigenic peptides from the cytosol to the lumen of the endoplasmic reticulum (ER) is an essential process for presentation to cytotoxic T‐lymphocytes. The transporter associated with antigen processing (TAP) is responsible for the intracellular translocation of peptides across the membrane of the ER. Efficient assembly of MHC‐peptide complex requires the formation of a macromolecular transport and chaperone complex composed of TAP, tapasin and MHC class I molecules. Therefore, structure and function of TAP is important for the understanding of the immune surveillance.

Programmed cell death protein 4 (PDCD4) acts as a tumor suppressor in neuroendocrine tumor cells.

Abstract: PDCD4 is a new tumor suppressor gene. In the current study, we show that overexpression of PDCD4 in carcinoid cells results in inhibition of cell proliferation. This is most likely caused by a PDCD4‐induced downregulation of carbonic anhydrase type II which catalyzes the production of bicarbonate, a fundamental substrate for many cellular pathways.