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Tensins 123
- 1. TENSINS IN HEALTH AND DISEASE MODERATOR: DR. MAHANTACHAR PRESENTER: DR. SPOORTHY
- 2. INTRODUCTION
- 3. The organs of multi cellular organisms consist of cells embedded in extra cellular matrix which execute specialist functions. During organogenesis, cells need to migrate to appropriate topographical location and stay there in order to maintain tissue integrity Following injury, cells need to migrate to damaged areas and undergo remodelling into the normal structure as a part of tissue repair
- 10. The correct assembly of the ECM is necessary for: 1. Direct morphogenesis during development 2. Maintenance of tissue integrity and patterning in the adult 3. Initiating cell signalling and events inside the cell
- 13. The attachment of the cell to the matrix takes place by the formation of cell adhesion complexes These include intergrins and many cytoplasmic proteins These adhesion complexes attach to the actin cytoskeleton and help in attachment to the ECM Thus they help the cell to endure pulling forces without being ripped off the ECM
- 14. INTEGRINS
- 15. Cell adhesion to the ECM is mediated predominantly by the integrin family of heterodimeric transmembrane glycoproteins Upon ligand binding, integrins activate signal transduction pathways: The presence of integrins allows rapid and flexible responses to the events at cell surfaces
- 19. FOCAL ADHESIONS
- 20. Focal adhesions are specialized regions of plasma membrane where cells attaches to the extracellular matrix (ECM) These are large molecular complexes which are generated following interactions of integrins with ECM Integrins help in their clustering at the site of action
- 21. 60 Focal Adhesion proteins are identified in vertebrates 1. Talin 2. Vinculin 3. Tensin 4. Paxillin 5. Focal adhesion kinase( FAK) 6. Protein kinase C 7. Src kinase
- 22. This supramolecular linkage appears to be critical in cytoskeleton organization and signal transduction, It is believed to participate in many biological events cell adhesion, migration, differentiation, proliferation, tissue development genesis of diseases
- 24. The life cycle of focal adhesions can be viewed:Assembly Maturation Disassembly
- 28. Tensin proteins are localised at focal adhesions These are among the molecules which act as molecular bridges between integrins and actin cytoskeleton The tensin family members play a pivotal role in key cellular processes including 1. Adhesion 2. Migration 3. Proliferation 4. Differentiation 5. Apoptosis
- 29. WHAT IS TENSIN?
- 30. Tensin is a 220Kd Cytoplasmic phosphoprotein It is localised in specialised areas of plasma membrane called FOCAL ADHESIONS Evidence is now emerging that suggests Tensin is an important component linking the ECM, Actin cytoskeleton and Signal transduction
- 31. Identified in mid-1980s as an actin binding contaminant of an Vinculin preparation Initially characterised on chicken tensin protein using: 1. SDS-PAGE 2. Dynamic light scattering 3. Gel filtration
- 32. Several groups reported that Tensin was localised to the ends of actin stress fibres at focal adhesions Tensin is phosphorylated on Tyrosine, Serine, Threonine residues Tyrosine phosphorylation of tensin is induced by 1. ECM attachment 2. PDGF treatment 3. Thrombin or Angiotensin 4. Oncogenes transformation: v-Src and Bcr/Abl
- 34. The Tensin gene family consists of 4 members 1. Tensin 1: TNS1 2. Tensin 2: TENC1 3. Tensin 3: TNS3 4. Tensin 4: TNS4 These genes encode multi domained cytoplasmic proteins
- 35. As a general structure of the Tensin family protein comprises of two domains. These domains bind to cytoskeletal proteins and signal transduction components. The Tensins have each been linked to multiple upstream and downstream signalling factors
- 37. The four members of the human tensin family display high sequence homology at the protein level All four tensins display significant homology at their C- terminus Tensins 1-3 have a high homologous N and C termini Divergent at their central regions
- 38. N- TERMINUS N-teminus comprises of: 1. Two actin- binding domains(ABD) 2. Region of homology to protein and lipid phosphatase(PTEN)
- 39. C-TERMINUS C-Terminus contains 1. Src-homology 2: SH2 domain 2. Phosphotyrosine binding domain: PTB
- 41. TENSIN 1
- 42. GENE Chromosome 2q 35-36 33 exons 1735 amino acid protein 185 KDa molecular mass Heart, skeletal muscles, kidney, lung
- 43. TEN1 has actin binding capabilities through distinct binding sites in the ABD Extra actin binding site is also present in the central region residues This central region shares high sequence homology to INSERTIN
- 44. C-TERMINUS: Tensin 1 contains SH2 regions similar to Tumour suppressor phosphatase and Tensin homologue (PTEN) This SH2 domain allows Tensin1 to bind to phosphorylated tyrosine residues on proteins such as 1. FAK 2. DLC 1 3. PI3 KINASE At both N and C termini, there are FAB domains which are required for localisation to focal adhesions
- 45. The PTB domains are known for binding phosphorylated tyrosine. Is is through this domain that Tensin 1 binds to the cytoplasmic tail of beta integrins independent of phosphorylation
- 47. TENSIN 2
- 48. GENE Chromosome 12q13 29 exons 1285 amino acid proteins 170KDa molecular weight
- 49. PROTEIN The central region of Tensin 2 is proline rich and can potentially act as a binding site for proteins containing Src homology(SH3) or WW domains This is the only member of Tensin family that has protein kinase C domain at N terminus
- 50. TENSIN 3
- 51. TENSIN 3- GENE Chromosome 7p12.3 1445 amino acid protein Molecular mass of 155KDa Placenta, Kidney
- 52. TENSIN 3-PROTEIN Tensin 3 contains 32 tyrosine residues, 13 of which are predicted to be potential sites of phosphorylation and possible candidates for signal transduction
- 53. TENSIN 4
- 54. TENSIN 4-GENE CTEN gene maps on chromosome 17q21.2 715 Amino acid protein 77KDa molecular mass Placenta, prostate tissues
- 55. CTEN- PROTEIN CTEN shows a high sequence homology at C terminus but it does not possess the N terminus This suggests that CTEN still contains the signalling component of other Tensin but lacks actin binding capabilities Thus it plays a novel role in cellular processes CTEN has also been detected in the nucleus
- 58. Exact individual role of Tensins are debatable In some circumstances can be tissue dependent Tensins role in health includes: 1. Cell adhesion and motility 2. Cell survival 3. Apoptosis
- 59. ROLE OF CELL MIGRATION
- 60. Cell migration is a complex process that is essential to the diverse range of organisms In humans it is required for a number of physiological events and several pathological conditions The exact role of individual tensins are tissue dependent They are important in: 1. Stabilising cell adhesion 2. Regulating cell motility
- 61. PHYSIOLOGICAL EVENTS: Gastrulation Wound healing PATHOLOGICAL CONDITIONS Vascular diseases Osteoporosis Rheumatoid Arthritis CANCER
- 63. TENSIN 2 BINDS TO DLC 1 INHIBITS Rho-GTPase activity PROMOTES CELL MOTILITY
- 65. CTEN EGFR SIGNALLING TENSIN 3 DECREASES CTEN levels increase TENSIN switch Lacks N terminus NO BIND ACTIN STRESS FIBRES CELL MIGRATION
- 68. Tensin Separates SH2/PTB from ABD Less of PI 3 mediated signal CASPASE 3 Integrins become detached
- 70. Two main pathways 1. EGFR signalling pathway 2. Stat3 pathwathy Other factors have been reported to regulate Tensin proteins ECM PDGF Thrombin Angiotensin Bcr/Abl
- 71. Stat 3 Signalling Stat 3 signalling has been implicated as one of the upstream pathways regulating expression of Cten Stat 3 dependent over expression of Cten has been shown to disrupt cell adhesion and induce motility IL-6 has been shown to induce Cten via Stat 3
- 72. EGFR SIGNALLING Activation of EGFR signalling through EGF stimulation leads to: Tensin switch which with a decrease in Tensin 3 levels and an increase in CTEN The other tensins remain unaffected
- 74. The signalling pathway between EGFR and Tensin 3 and CTEN mostly involves KRAS/BRAF/ Mitogen activated protein kinase pathway Hence a mutation activation of KRAS and BRAF has been shown to up regulate Cten
- 76. Several studies have highlighted alterations in the expression of the human tensins in the cancer Tensins are concerned with cell motility their involvement in carcinogenesis particularly the promotion of metastasis Role for Tensins in cancer has been most rigorously investigated for Tensin 3 and Cten Evidence clearly implicates each family member in disease progression
- 77. TENSIN 1 Expression of Tensin 1 was shown to be down- regulated in prostate and breast cancer cell lines Tensin 1 expression was induced in response to Resveratrol treatment in tensin negative or deficient erythroleukemic and breast cancer cell lines
- 78. TENSIN 2 Tensins 2 has been reported to be down regulated in cancers of kidney and lung Investigations in HCC revealed Tensin 2 to be over expressed in 46% of tumours compared to normal liver tissue
- 79. TENSIN 3 Tensin 3 reduced expression has been reported in the tumours of Thyroid, Kidney and Breast Forced expression of Tensin 3 led to decreased colony formation This supports the role of TEN 3 as a tumour suppressor This is consistent with the Tensin 3 ability to decrease cell migration in normal mammary epithelial cells
- 80. CTEN CTEN was originally identified as a tumour suppressor gene in prostate cancer It is also down regulated in tumours of kidney CTEN is normally not present in all tissues Elevated levels can be seen in tumours of Lung, Thymus, Colon, Breast and Pancreas This suggests that in these tumours it acts as an oncogene Greater levels are associated with advanced disease stage and metastasis
- 81. IHC analysis of normal and cancerous prostate tissue was done It revealed that CTEN expression was inversely correlated with prostate tumour grade CTEN is frequently over expressed in gastric cancer Tumours with higher CTEN expression displayed more aggressive behaviour
- 82. In breast cancer, CTEN expression has been shown to stimulate cell motility. CTEN was shown to be associated with tumour size, grade, nodal involvement and poor nottingham prognostic index CTEN expression was correlated with evidence of tumour progression in lung cancer and thymoma with metastasis to lymph node
- 83. CTEN interaction with beta- catenin also contributes to enhancing tumorigenicity Beta- catenin is found in adherent junctions, cytoplasm and nucleus In the nucleus, beta-catenin binds to TCF/LEF forming a transcriptional complex that turns on gene expression CTEN is reported to interact with beta-catenin only in the nucleus
- 85. The renal histology of Tensin null mice revealed a typical triad of human nephronophthisis 1. Tubular membrane disruption 2. Tubular ectasia 3. Interstitial inflammation Hence Tensin null mouse could be used as an animal model for human nephronophthisis or similar renal diseases
- 86. Tensins contains tri nucleotide CAG repeats Expansion of CAG repeats has been detected in many proteins that are relevant to inherited diseases It will be intriguing to know whether an expansion of tensin CAG repeats plays an important role in pathogenesis of human diseases
- 87. Analysis of Tensin expressions in human cancer patients and cell lines speculate that the loss of tensin expression may be an early event involved in cell transformation Tensins and its down streaming signalling molecules could be bio markers or therapeutic interventions in cancer
- 88. CONCLUSION
- 89. Tensins are emerging at the forefront as regulators of cell migration They have the ability to both bind actin cytoskeleton and mediate signal transduction events at focal adhesion regions This conveys their importance in regulating cell adhesion and migratory processes The Tensin family’s role in regulation of cell migration in turn extends to their involvement in cancer metastasis
- 90. The targeting of the cell migratory machinery by using tensin family gene presents an interesting therapeutic target for anti cancer therapies. Further characterisation may provide alternative targets in areas of need including EGFR inhibitor resistant tumours Over come problems associated with toxic anti- Src therapies under development
- 91. REFRENCES 1. 23 Recent advances in histopathology 2. Lo SH. Tensin. The international journal of biochemistry & cell biology. 2004 Jan 1;36(1):31-4. 3. Huaiyang CH, ISHII A, Wai-Keung WO, CHEN LB, LO SH. Molecular characterization of human tensin. Biochemical Journal. 2000 Oct 15;351(2):403-11. 4. Martuszewska D, Ljungberg B, Johansson M, Landberg G, Oslakovic C, Dahlbäck B, Hafizi S. Tensin3 is a negative regulator of cell migration and all four Tensin family members are downregulated in human kidney cancer. PloS one. 2009 Feb 4;4(2):e4350. 5. Liao YC, Lo SH. Deleted in liver cancer-1 (DLC-1): a tumor suppressor not just for liver. The international journal of biochemistry & cell biology. 2008 Jan 1;40(5):843- 7.
- 92. THANK YOU
Editor's Notes
- The process of organogenesis, organ homeostasis, tissue repair require modulation of cell adhesion to enable cell motitliy and maintain cell stasis let us now explore at the molecular level what causes this cell migration how cells modulate different factors for their functions.
- since the first time the word cell was used by robert hooke many scientists have tried to decipher this tiny unit of our body and with the advances in the technology we are still having new break throughs every single day. this is a picture the depicting the parts of an animal cell. for our topic we are mainly concerned with the internal environment of the cell and the external environment which are clearly demarcated by the cell membrane and how they interact with each other
- the structure of cell membrane is explained by the fluid mosaic model. in addition to the lipid bilayer, it also contains a number of proteins. these membrane proteins allow for many of the interactions that occurs between cells. these membrane proteins are broken into different classifications like1. integral proteins 2. lipid bound proteins 3. peripheral proteins INTEGRAL PROTEINS: these proteins are embedded within the lipid bi layer. these proteins float freely in the lipid bi layer. these integral proteins are usually trans membrane proteins, extending through the lipid bilayer. one end is in contact the interior of the cell and other touches the exterior. as a result of their structure these are the only class of proteins that can perform functions both inside and outside of the cell
- 3 types of protein fibres in the cytoskeleton: micro filaments, intermediate filaments and microtubules they support the plasma membrane and gives the cell an overall shape.
- micro filaments: they are found in the region of the cytoplasm at the edge of the cell. actin and myosin pair with each other and help in the movement of the cell, like leukocyte movement intermediate filaments: they are keratin, neuro filamentsand nuclear lamimins they play a structural role of the cell, maintaining the shape microtubules: tubules they are hollow. they help the cell to resist the compression forces
- The Extra cellular matrix is composed of many different proteins: Collagen, Laminin, Fibronectin Polysaccharides: GAG class These covalently attached to proteins to form proteoglycans These molecules interact with each other to form macromolecular complex.
- The polysaccharides of the ECM form a highly hydrated gel, which resists compressive forces. Permits the diffusion of smaller molecules through tissues The fibrous proteins are embedded within this gel helps to determine the structural and adhesive properties of the ECM
- Cell adhesions can be viewed as intercellular adhesions or cell-matrix adhesions. the former is maintained through intercellular junctions. while the cell matrix adhesions is maintained by focal adhesions
- integrins couple the ECM to the cytoskeleton inside the cell. which ligand in the ECM can bind to the I is decided by the alpha and beta sub units. these include talon, vincullin, paxillin Integrins lie at the heart of many cellular biological processes
- signal platelets to initiate an interaction with the coagulation factors. the prominent function of interns is seen in the molecule gp11a/3b, an integral on the surface of blood platelets, responsible for attachment to the fibrin for developing blood clot. gp11a/3b, changes shape allowing it to bind to fibrin and other blood components
- STRUCTURE OF INTEGRIN heterodimers. i.e they have 2 sub units. alpha and beta.parts include: head, thigh, transmembrane helices and a cytoplasm tail. these interns hence serve as a link between two networks across the plasma membrane i.e the extra cellular matrix and the intracellular actin- filamentous system
- initial binding of integrins to EC ligand will activate the integrins. this causes conformational change in the integrins. this causes recruitment of adapter proteins and cytoplasmic changes will result in clustering of other integrins at the site. this is called positive feed back loop. this results in the formation of early focal adhesions called nascent adhesions. where ECM and cytoskeleton are first linked. these may undergo dissolution or following Rac dependent actomyosin contraction which expands to form focal complexes. rapid formation and dissolution of nascent adhesions and focal complexes are a feature of migratory cells. but these may also progress to mature focal adhesions which are characterised by contraction of myosin and and recruitment of molecules such as vinculin this exerts tension on focal adhesions and stabilises the connection net wen actin cytoskeleton
- these may undergo dissolution or following Rac dependent actomyosin contraction which expands to form focal complexes. rapid formation and dissolution of nascent adhesions and focal complexes are a feature of migratory cells. but these may also progress to mature focal adhesions which are characterised by contraction of myosin and and recruitment of molecules such as vinculin this exerts tension on focal adhesions and stabilises the connection net wen actin cytoskeleton
- in stable non motile cells, maturation of focal adhesions is completed by the formation of large actin stress fibres and recruitment of tensins. this stage is called fibrillar adhesions. disassembly of focal adhesions is mediated through unbiquitination and proteosomal degradation of talin or degradation of ECM thereby removing the signals for intern activation
- The oncogenic focal adhesion proteins contribute to multiple facets of cancer Increased cell proliferation, Resistance to apoptosis, Elevated cell motility Invasion
- consistent with the capping activity
- Tennis 1,2,3 are large and multi-domained proteins cten is a shortened protein which is half the length of the larger tensions
- insertin is an actin capping protein which retards globular actin polymerisation
- identified as a protein related to tennis 1 through homologous nand c terminals with 60% and 67% sequence similarity similar tissue distribution as 1
- seen in most of the tissues particular prevalence in placenta and kidney spleen lund askeletal muscle and heart showing lower expression
- The aberrant invasive migration of cells away from the primary tumour ultimately leads to the formation of distant metastasis
- The recruitment of tensins to focal adhesions is dependent on the stage of maturation of focal adhesions. tennis 2 is recruited in early focal adhesions through its PTB domain.
- Tensin 1 and 3 are found in mature form of focal adhesions known as fibrillar adhesions. tension 1 has DLC-1 binding sites in SH2 domain. tenisn 3 has DLC-1 binding sires in ABD domain.tensin 3 has results in increased Rho gtpase activity causing it to inhibit cell motility. so in our tensins we have tension 2 which promotes cell motility and tension 3 which inhibits cell motility
- These effects are mediated through phosphorylation of tensin residues
- indicating a tumour suppressor activity
- similar to tennis 1 and 2 tennis 3 generally appears to have tumour suppressive role
- high tumour expression of cten had poor prognosis in comparison to those expressing low levels and also had increased chances of metastasis
- Over the past decade, the emphasis in anticancer drug discovery has shifted from an empirical approach, characterized by random screening of a variety of natural and synthetic compounds using high throughput cell-based cytotoxicity assays, to a more rational and mechanistic, target-based approach [1]. The goal of this new target-based approach is to improve the efficacy and selectivity of cancer treatment by developing agents that specifically block the pathogenic mechanisms that account for malignant transformation. This new approach reflects our rapidly expanding knowledge of the pathogenesis of a variety of forms of cancer at the molecular level, providing new targets for drug discovery. Drugs that validate the molecularly targeted approach to anticancer drug development include tretinoin (all-trans-retinoic acid), which targets the PML-RARα fusion protein in acute promyelocytic leukemia (APL) [4, 5], and imatinib mesylate (STI571, Gleevec™), which targets the BCR-ABL fusion protein in chronic myelogenous leukemia [6, 7] and the mutated KIT receptor in gastrointestinal stromal tumors (GIST) [8]