1 / 35

Wallace McKeehan Center for Cancer & Stem Cell Biology

II. II. F. F. III. III. HSPG. SNT1. Grb2. SHP2. sos. sos. Ras. Grb2. SNT1. AKT. PI3K. Raf. PKC. PLC- g. PLC- g. MEK. ?. ERK1/2. Forty Years of FGF: Regulator of Cellular & Metabolic Homeostasis. Wallace McKeehan Center for Cancer & Stem Cell Biology. The Klothos.

willow
Download Presentation

Wallace McKeehan Center for Cancer & Stem Cell Biology

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. II II F F III III HSPG SNT1 Grb2 SHP2 sos sos Ras Grb2 SNT1 AKT PI3K Raf PKC PLC-g PLC-g MEK ? ERK1/2 Forty Years of FGF: Regulator of Cellular & Metabolic Homeostasis Wallace McKeehan Center for Cancer & Stem Cell Biology The Klothos

  2. Gordon Sato, The Mangrove Man Gospodarowicz D, Jones KL, Sato G. 1974 Purification of a growth factor for ovarian cells from bovine pituitary glands. PNAS 71:2295 Gospodarowicz D. 1975 Purification of a fibroblast growth factor from bovine pituitary. JBC 250:2515 Maciag T, Mehlman T, Friesel R, Schreiber AB. 1984. Heparin binds endothelial cell growth factor, the principal endothelial cell mitogen in bovine brain. Science 31;225:932. Gospodarowicz D, Cheng J, Lui GM, Baird A, Böhlen P. 1984. Isolation of brain fibroblast growth factor by heparin-Sepharose affinity chromatography: identity with pituitary fibroblast growth factor. PNAS 81:6963. 1985-1986: Cloning of FGF1 and FGF2 1989-1993: Cloning and diversity of FGFR tyrosine kinases via splice variants 1993-2002: Structure of FGFR kinases and interaction with heparan sulfate

  3. FGF1 FGF2 FGF3 FGF4 FGF5 FGF11 FGF12 FGF16 FGF17 FGF18 Structure, Mechanism of Assembly & Signaling of the FGFR Complex Continued Discovery Through Homology & Genome Sequencing: 22 FGF Homologues 18 High Affinity FGFR Activators FGF9 FGF8 FGF6 FGF7 FGF10 Four Transmembrane Tyrosine Kinase Receptors Numerous Splice Variants: FGFR1-4 Blue: Positive Charge Red: Negative Charge White: Neutral FGF21 FGF23 FGF20 FGF22 FGF19

  4. II II II II II II F F F F III III III III III III II II II F II III F III III F III + + + + + + Xu…McKeehan, JBC 1992 Kan…McKeehan, Science 1993 Yayon…Ornitz, Cell 1991 Pantoliano…Sisk Biochem. 1994 II II F F F F III III F F Spivak-Kroizman…Schlessinger 1994 Kan…McKeehan, JBC 1996 Venkataraman…Sasisekharan, PNAS 1996 Moy…Powers, Biochem. 1997 II II F F III III Plotnikov…Mohammadi, Cell 1999 McKeehan, Wang, Kan, PNAR 1998 DiGabriele…Hendrickson, Nature 1998

  5. II II F F III III Kan…McKeehan, JBC 1996 McKeehan, Wang, Kan, PNAR 1998 Schlessinger….Mohammadi, Cell 2000 Conformational Model of Control and Activation (Derepression) of the FGFR Complex Pre-existent unliganded symmetric complex of 2:2 FGFR-HS Conformational maintenance of dependence on FGF for kinase derepression by transphorylation by heparan sulfate in the ectodomain Conformational transmembrane communication between ecto and intracellular domains Pelligrini…..Blundell, Nature 2000

  6. II II F F III III HSPG SNT1 sos Grb2 sos Ras Grb2 SNT1 AKT PI3K SHP2 Raf PKC MEK PLC-g PLC-g ? ERK1/2 How Can FGF-dependent Change Outside Be Transmitted Across the Membrane? Next Generation: Are Intracellular Substrates/Adaptors Organized and Waiting for Conformational Derepression? F F

  7. FGF HS proteoglycan COO- CH2OX O O Core protein Linker COO- HS FGFR OH OX O O NY OX n IdoA/GlcA GlcN Role of Heparan Sulfate 2-O-, 2-N-, 3-O- and 6-O-sulfates, 2-N-acetyl and epimerization 1. Matrix and Membrane: Reservoir, Stabilizer, FGF Delivery System 2. Integral Component of the FGFR Complex: Specificity for FGF Assembly of the oligomeric complex Negative restriction of the unliganded dimeric complex Stabilization of the derepressed, ligand-activated complex

  8. Is There Structural Specificity in Heparan Sulfate Beyond Variations in Charge Density? 7,8-S-OctaF7, which has anticoagulant activity and likely the motif containing a 3-O sulfate, specifically supports high affinity FGF7 binding and mitogenesis in cells expressing FGFR2IIIb. It failed to support high affinity FGF1 binding and mitogenesis. Purification of an undersulfated 7,8-S-octasaccharide mixture (7,8-S-OctaF7) by affinity chromatography with FGF7 Hypothetical deduced structure of 7,8-S-OctaF7. The x, y or z could be a sulfate, preferably 6-O-sulfate with the other two as hydrogens (8-S), or x, y and z may all be hydrogens (7-S).

  9. The Klotho Co-factors: single pass transmembrane proteins involved in endocrine FGF activities SS TM Similarities to Heparan Sulfate (HS): Independent binding to eFGFs or FGFR Direct participation in the FGFR complex with both FGF and FGFR Determinants of specificity for eFGFs Are HS and Klothos at play in the same FGFR complex? Do HS or Klotho co-factors alter quality of the FGFR signal? Does the protein core of HS or the intracellular domain of HS or Klothos play a role in signaling? αKlotho (1014aa) N C αKl1 αKl2 βKlotho (1043aa) N C βKl1 βKl2

  10. Development and Adult Homeostasis Modes of Function: Autocrine, Paracrine, Endocrine Development: Autocrine and Paracrine Control: Changing, Short-lived Cell Autonomy, Transcription rate-limiting Adult Tissue Homeostasis: Largely Paracrine Control: Partition of FGF and FGFR between Cells, Activity, not transcription rate-limiting Autocrine is Pathological Intracrine (FGF, FGFR or FGF-FGFR)?

  11. Is there tissue and target cell specificity among the 18 FGFs and the 4 tyrosine kinase receptors and variants? Specificity set by: 1. Paracrine partition of FGF and the FGFR complex between cells. 2. Endocrine partition of FGF and FGFR complex between organs. 3. Cell-specific co-receptors HS and klothos. Do different FGFR isotypes have exhibit different signaling endpoints in the same context?

  12. Epithelium FGR2b FGF 7 FGF10 FGF9 Heparan sulfate FGFR3 Stroma Subversion and Autocrine Switch of Canonical Matrix-controlled Short Range Paracrine FGF Signaling Results in Pathologies Loss of R2b EctopicFGFR1 Autocrine Switch Cancer Feng, Wang, Matsubara, Kan, McKeehan. Fibroblast growth factor receptor 2 limits and receptor 1 accelerates tumorigenicity of prostate epithelial cells. Cancer Res. 1997

  13. Overlapping and Distinct TyrP Targets Between: FGFR2IIIb FGFR1IIIc RSK2 Fyn FAK ShcA P85α P85β Paxillin PTPN18 CDK2 (Tyr15) Emerin ZRF1 LAP2 SAP102 FGFR SHP2 SHIP2 ERK2 IRS4 FRS2 PLCλ growth/tumor suppression, nuclear-cytosol interplay, cell structural maintenance growth promotion, cell survival, adhesion, motility Luo et al. Novel phosphotyrosine targets of FGFR2IIIb signaling. Cell Signal 2009

  14. Hepatocyte FGFR4 Hepatocyte Cyp7a Cholesterol to Bile Acids 2000: First implication of FGF signaling in regulating metabolic circuits: Cholesterol/bile acid homeostasis A B 5 +/+ 90 +/+ 80 -/- -/- 70 4 60 50 3 40 30 2 20 10 1 0 0 24 48 72 96 120 144 168 0 24 48 72 96 120 144 168 Time after partial hepatectomy (hr) Yu, Wang, Kan…McKeehan. Elevated cholesterol metabolism and bile acid synthesis in mice lacking membrane tyrosine kinase receptor FGFR4. JBC 2000

  15. Evolution of the Endocrine FGFS Early 2000: FGFR4 involved in cholesterol/bile acid metabolism. Yu et al. JBC 2000 Late 2000: Mutation in FGF23 involved in mineral metabolism. ADHR Consortium, Nat Genetics 2000 2002: FGF19 impacts metabolic rate and adiposity. Tomlinson et al., Endocrinol 2002 2005: FGF21 is a metabolic regulator Kharitonenkov et al., JCI 2005 2005: Klothos impact metabolism & FGF signaling The FGF15/19-FGFR4 Axis: An enterohepatic regulator of cholesterol/bile acid homeostasis Inagaki et al., Cell Metab. 2005

  16. Steroids FGF21 effectors have expanded! Steroidreceptors Paracrine FGFsFGF7,9,10, others heparan sulfates FGFR2IIIbFGFR3 Cellularcompartmentalhomeostasis D.D. Moore, Science 2007

  17. Hepatocyte FGFR4 Has Multiple Effects Beyond Hepatic Cholesterol/Bile Acid Metabolism 1. Limits extent of toxic liver injury and fibrosis 2. Modulates hepatic lipid and glucose metabolism 3. Supports fatty liver in obesity or starvation 4. Reported as both hepatoma promoter and hepatoma suppressor Which effects beyond bile acid metabolism occur directly within hepatocytes due to FGF19-FGFR4-bklotho signaling is unclear.

  18. R4 R1 R4 Hepatoma cells Hepatocytes Resident hepatocyte FGFR4 limits hepatocarcinogenesis while ectopic hepatocyte FGFR1 accelerates it Huang, et al. Ectopic activity of fibroblast growth factor receptor 1 in hepatocytes accelerates hepatocarcinogenesis by driving proliferation and vascular endothelial growth factor-induced angiogenesis. Cancer Res 2006 Huang, et al. Resident hepatocyte fibroblast growth factor receptor 4 limits hepatocarcinogenesis. Mol Carcinog. 2008 Ectopic FGFR1 in Hepatocytes WT R1TG WT R1TG FGFR4 Knockout

  19. bklotho (KLB) is reduced in human & mouse hepatomas Without comparable changes in FGFR4 8 Human SIB-CleanEX DbGSE7307;2109 6 Normal Liver 4 Hepatoma 2.5 2 Relative Expression KLB FGFR4-/-/FGFR4ecto P=0.0018 0 40 FGFR4-/- 2 -2 30 Apoptosis (A550 x 100) 1.5 Relative Expression of KLB -4 20 1 -6 10 0.5 -8 0 0 0.04 0.2 1 5 KLB construct 0 Normal liver FGFR4-/- Hepatoma Hepatoma Restoration of KLB and FGFR4 to KLB- and FGFR4-deficient mouse hepatoma cells induces apoptosis

  20. HEK293 KLB Constitutive(c) FGFR4 Induced(i) cKLB + iFGFR4 iR4 cKLB F19 8 iR4 cKLB F1 iR1 cKLB F19 iR1 cKLB F1 iR4 cKLB iR4 F1 iR1 cKLB 293 cKLB iR4 iR4 F19 (R4) cKLB 6 Cell Number x 10-5 4 2 0 KLB drops in liver after partial hepatectomy and recovers during restoration Relative Expression Day after partial hepatectomy KLB partners with FGFR4 to inhibit cell population growth via apoptosis Apoptosis induced by the FGFR1/4-KLB pair is enhanced by either FGF1 or FGF19 FGF21 is specific for the FGFR1-KLB partner

  21. KLB can confer growth controlling, anti-tumorigenic pro-apoptotic activity on both FGFR4 and FGFR1 signaling complexes. This is in addition to the role of KLB in conferring high affinity of FGFR4 for endocrine FGF19 and FGFR1 for both FGF19 and FGF21. Direction of pro-apoptotic signaling is likely through KLB-dependent abrogation of anti-apoptotic AKT and mTOR pathways How might this KLB-dependent redirection occur? KLB-dependent abrogation of anti-apoptotic AKT and mTOR pathways?

  22. II II II II cF cF III III F F Klothos III III HSPG HSPG SNT1 SNT1 sos Grb2 Grb2 sos Ras Grb2 SNT1 AKT PI3K SHP2 SHP2 sos sos Ras Grb2 SNT1 AKT Raf PI3K PKC Raf MEK PKC PLC-g PLC-g PLC-g PLC-g ? MEK ERK1/2 ? ERK1/2 Canonical(c) FGF Paracrine/Autocrine Signaling Endocrine(e) FGF Signaling cF cF eF F1 Local Local Distal Local Cellular Homeostasis (Development & Adult) Growth, migration, morphogenesis Promotion of Tumor Phenotype Metabolic Homeostasis Inhibition of growth, pro-cell death Tumor Suppression

  23. Net KLB-directed anti-growth and anti-tumor effects is consistent with primary function of eFGFs in control of metabolic homeostasis. Targeting hepatic FGFR4 will disrupt roles in metabolic homeostasis, particularly bile acid metabolism, and may have tumor-promoting effects. ******************************************************************************************************* Nicholes et al. A mouse model of hepatocellular carcinoma: ectopic expression of fibroblast growth factor 19 in skeletal muscle of transgenic mice. Am J Pathol 2002 Desnoyers et al. Targeting FGF19 inhibits tumor growth in colon cancer xenograft and FGF19 transgenic hepatocellular carcinoma models. Oncogene 2008 French et al. Targeting FGFR4 inhibits hepatocellular carcinoma in preclinical mouse models. PLoS One 2012 ******************************************************************************************************* Mellor. Targeted inhibition of the FGF19-FGFR4 pathway in hepatocellular carcinoma; translational safety considerations. Liver Int. 2014

  24. FGF21 Ileal FGF19 is under post-prandial bile acid-control. FGF21 is significant only under conditions of metabolic extremes as starvation and obesity and a wide range of conditions causing systemic and tissue stress. Liver has been the major notable source of FGF21 during metabolic stress. FGF21 binds and activates the FGFR1-KLB complex, but not the FGFR4-KLB complex. FGFR1 is the major FGFR isotype in adipocytes where KLB is notably expressed. (FGFR1 is not significant in hepatocytes where FGFR4-KLB dominates) FGF19 binds and activates both FGFR4- and FGFR1-KLB complexes ************************************************************************************ Are there non-hepatic sources of FGF21? Is adipocyte FGFR1-KLB the primary direct target of FGF21 during metabolic stress? Does adipocyte FGFR1 account for the beneficial effects of FGF21 and FGF19 on obesity and diabetes? What is the role of FGF21 activation of adipocyte FGFR1-KLB in alleviation of stress?

  25. Are there non-hepatic sources of FGF21? FGF21 is induced by diverse sources of stress in diverse tissues Liver: Starvation, obesity, chemical and genetic disruptions (LKB1, p53, MST1/2, SAV1 and PTEN), loss of liver mass (partial hepatectomy), hepatitis, fatty liver, cirrhosis, hepatoma. Yang et al. BMC Gastroenterology 2014 ************************************************************************ Skeletal, heart, gastrocnemius muscle: Mitochondrial respiratory, defects, autophagy/mitophagy defects, chronic exercise, Fe-S cluster defect. Suomalainen et al. Lancet Neurol 2011 ; Yang et al. Clin Endocrinol 2011 ; Cuevas-Ramos et al. PLoS One 2012 ; Kim et al. Nat Med 2013 ; Crooks etal. Hum Mol Genet 2014 ********************************************************** Diverse Transcriptional Regulators of FGF21: PPARα/γ, RAR, ChREBP, SREBP1c, LXR, STAT3/5, p53, ATF4, MIST1 Brown fat: Cold stress, fetal-neonatal transition. Hondares et al. Cell Metab 2010 ; J Biol Chem 2011 *********************************************************** Pancreas: Pancreatis. Johnson et al. Gastroenterology 2009 ; Am J Physiol Endocrinol Metab. 2014 *********************************************************** Is adipocyte FGFR1-KLB the primary direct target of FGF21 during metabolic stress?

  26. Ablation of FGFR1 in Adipocytes Poising transcriptional activity of hepatic lipogenic genes without much effect on adipocyte genes. Indirect increase in hepatic steatosis concurrent with increase in hepatic lipogenic genes without much effect on adipocyte gene expression. Elevation of triglyceride and NEFA without effect on glucose or ketone bodies under starvation conditions concurrent with an increase in adipocyte lipase activity.

  27. FGF21/FGFR1-mediated adipo-hepatic communication Under stress FGF21 is induced in liver. FGF21 activates adipose FGFR1 that imposes restrictions on adipocyte lipolysis and indirectly hepatic lipogenesis. This attenuates extent of potentially damaging hepatic steatosis that often occurs during hepatic stress caused by starvation. Lipolysis and lipogenesis are normally tightly coupled inversely with each other and to glucose and ketone body metabolism. Uncoupling may mete out and extend lipid reserves for neural fuels (glucose and ketone bodies) during metabolic extremes and other conditions causing hepatic stress. This mechanism may underlie the beneficial effects of FGF21 (and in part FGF19) under metabolic extremes as well as other sources of hepatic stress. Is adipocyte FGFR1 the direct mediator of the beneficial effects of FGF21 under stress of obesity and diabetes?

  28. Adipocyte-specific ablation of FGFR1 indicates that adipocytes via FGFR1 account for effects of FGF21 and likely the effects of FGF19 • Yang et al. Control of lipid metabolism by adipocyte FGFR1-mediated adipohepatic communication during hepatic stress. Nutr Metab (Lond). 2012 • Adams & Yang et al. The breadth of FGF21s metabolic actions are governed by FGFR1 in adipose tissue. Mol. Metab. 2012 • Foltz et al. Treating diabetes and obesity with an FGF21-mimetic antibody activating the βKlotho/FGFR1c receptor complex. Sci Transl Med. 2012

  29. Adipose FGFR1 deficiency abolishes weight loss, glucose and energy regulation by FGF21 in the obese Foltz et al. Treating diabetes and obesity with an FGF21-mimetic antibody activating the βKlotho/FGFR1c receptor complex. Sci Transl Med. 2012

  30. Adipose FGFR1 deficiency also abolishes weight loss, glucose and energy regulation by FGF19 in the obese "...while subtle differences exist in vitro, the metabolic effects in vivo of FGF19/FGF21 are indistinguishable, supporting a shared mechanism of action for these two hormones in the regulation of energy balance." Adams...Kharitonenkov PLos One 2012.

  31. FGFR4/KLB: Hepatic Cholesterol to Bile Acids Hepatic Lipogenesis Ileal FGF19 Conclusion During normal feeding ileal FGF19 is likely the normal coordinator of adipo-hepatic communication in the lipid metabolism domain. FGF21 is a hepatokine reserved to instruct adipocytes under conditions of metabolic extremes and other stress conditions sensed by the liver. Effects of FGF19/FGF21 on glucose metabolism are largely an indirect consequence of direct regulation of lipid metabolism in both liver and adipocytes in both normal and extreme conditions.

  32. FGFR4/KLB: Hepatic Cholesterol to Bile Acids Hepatic Lipogenesis Ileal FGF19 Conclusion During normal feeding ileal FGF19 is likely the normal coordinator of adipo-hepatic communication in the lipid metabolism domain. FGF21 is a hepatokine reserved to instruct adipocytes under conditions of metabolic extremes and other stress conditions sensed by the liver. Effects of FGF19/FGF21 on glucose metabolism are largely an indirect consequence of direct regulation of lipid metabolism in both liver and adipocytes in both normal and extreme conditions.

  33. Recap, Summary Overall

  34. Dr. Chundong YuProfessor BiologyXiamen University Dr. Xinqiang Huang Regulus Therapeutics Dr. Yongde LuoAssistant Professor IBT Texas A&M Chaofeng YangPostdoctoralUT Southwestern US Public Health Service grantsJohn S. Dunn Research Foundation Komen Breast Cancer Foundation Amgen & Eli Lilly Dr. Fen WangProfessor Center DirectorIBT Texas A&M

More Related