Historical perspective of FGF and related proteins
Fibroblast growth factors (FGFs) were first discovered in the 1970s as a mitogen that can stimulate the growth of fibroblasts. In 1974, the first growth factor was isolated from cultured fibroblasts. While the mitogenic activity was their first known function, FGFs have since been found to be involved in much more than just fibroblasts. FGFs are a large family made up of polypeptides that mediate cell proliferation, cell differentiation, angiogenesis, tumor growth, and wound healing. All FGFs share a common high affinity for heparin.
There are 22 members in the FGF family. FGF-1 and FGF-2 are the most well-known of them. Most of the FGFs mediate their biological processes such as wound healing by acting as extracellular proteins. These FGFs bind to and activate their transmembrane receptor, FGFR. FGF-1 is sometimes referred to as aFGF or acidic FGF and FGF-2 is referred to as bFGF or basic FGF. These two FGFs were given many names during the initial characterization of these proteins. This shows the significance of these proteins and the important role they play in many biological activities. FGF-1 and FGF-2 do not contain a signal peptide. It was first thought they were released from dead or dying cells in order to assist in the destruction of cell/tissue. It was later shown that low levels of FGF-2 were released from healthy cells. These FGFs can activate their own receptors via autocrine manner.
In 1983, it was first shown that heparin or sulfated glucosaminoglycan could activate the biological activity of acidic FGF. This pioneer work set the foundation for the discovery of acidic and basic FGF ability to bind to heparin. Several studies showed that acidic FGF eluted from heparin at 1.0M NaCl while basic FGF needed a higher concentration of salt ~1.6M NaCl to elute from heparin. In the late 1980's the few FGFs that had been discovered were separated into two classes: Class I which describes the acidic FGFs and Class II which describes the basic FGFs.
Subfamilies of FGF Based on Function
As mentioned earlier, there are currently 22 FGFs in the mammalian FGF family. Itoh and Ornitz have further classified these 22 FGFs into three separate sub families based on their function. These three sub families include: canonical, hormone like, and intracellular FGFs.
Canonical FGFs consist of FGF 1-10, 16-18, 20, and 22. This group of FGFs is very specific in that they only interact with their receptor at a high affinity. These proteins are secreted, bind, and then activate their receptors. These proteins are involved in cell proliferation and differentiation, and all have binding ability for heparin and heparan sulfate.
Hormone like FGFs include: FGF-15, FGF-19, FGF-21, and FGF-23. These FGFs have a lower affinity for heparin and can act on target cells in other areas of the body. To activate their receptor they require cell surface co-receptors, Klotho or βKlotho, which are expressed by target cells. FGF-15 and FGF-19 are secreted from the intestine upon food ingestion, FGF-21 is secreted from the liver, and FGF-23 is secreted from bone. Upon secretion, FGF-23 can suppress renal phosphate re-absorption and vitamin D production in the kidneys, which consequently lowers blood phosphate levels. FGF 21 is involved in adipose tissues and promotes lipolysis. FGF-21 is also involved in glucose uptake by increasing glucose transporter-1. FGF-15 and 19 interact with the liver to decrease bile acid synthesis.
Intracellular FGFs include FGFs 11-14. These FGFs do not interact with the FGF receptor. These FGFs have been shown to interact with the intracellular domains of the salt channels and with the neuronal mitogen-activated protein kinase scaffold protein, islet-brain-2.
Chemistry and broad functions of FGFs
FGF-1 is located on chromosome 5 and it is the most widely expressed isoform. It was first detected in and purified from bovine brain tissue extracts. FGF-1 lacks the N terminal peptide signal and therefore does not use the classical secretion pathway like many proteins; instead it uses a non-classical pathway. FGF-1 has been found in both embryonic and adult tissues and can stimulate proliferation of many different cell types including mesodermal cells. FGF-1 is known as the universal ligand since it can bind to all FGF receptors. One of the important functions of FGF-1 is angiogenesis. Angiogenesis is the process of creating new blood vessels from pre-existing vessels, and is obviously a vital process for the body. Angiogenesis occurs during the development of the vascular system during embryonic, fetal, and adolescent development. The function of FGF in angiogenesis is to stimulate the proliferation and differentiation of many different cells including endothelial and smooth muscle cells.
FGF-2 can be found on chromosome 4. FGF-2 has four isoforms that have identical cores. These four FGF-2 isoforms have a molecular weight of 18kDa, 21kDa, 22kDa, and 25kDa. The 18kDa FGF-2 isoform is cytoplasmic and like FGF-1, it plays a role in cell proliferation of many cells including the endothelial cells. The other isoforms of FGF-2 can be found in the nucleus and play a role in controlling transcription.
FGF-3 can be found on chromosome 11. Originally known as Int-2, FGF-3 is a 240 amino acid protein that is 40% homologous to FGF-1 and 2. The function of FGF-3 is not very well understood but, is thought to play a role in embryonic development. FGF-3 can remain in the cell and travel to the nucleus where they are involved in regulating gene transcription. FGF-3 is known to cause a skeletal disorder known as, "Michel aplasia". This disorder is recognized by complete loss of the inner ear structures. There are one mis-sense and two nonsense mutations in FGF-3 involved in this mutation.
FGF-4 is also found on chromosome 11 in a close proximity to the FGF-3. It plays an important role in limb regeneration and in the development of teeth. FGF-4 can be found in some adult tissue, but it is predominately found during embryonic development. FGF-4 is about 40% homologous to FGF-1 and 2 and contains 267 amino acids.
FGF-5 is expressed during embryonic development and can be found in adult tissues. FGF-5's main function is in embryonic development of the brain, muscle, and heart. In adult tissues FGF-5 and FGF-6 play a role in development of mesenchyme.
FGF-6 is also involved in wound healing and tissue regeneration. FGF6 share 30% homology with FGF-1 and is 70% homologous with FGF-5.
FGF-7 is one of the most well studied FGFs and was first known as keratinocyte growth factor. It is 35% homologous to other FGFs, and is the only FGF to show specificity to epithelial cells. Interestingly, FGF-7 has no angiongenic activity.
FGF8 is 30% homologous with other FGF family members. The FGF-8 gene is located on chromosome 10 and is found in the embryonic stage of development and plays an important role in outgrowth and patterning for limb development. Mutations in FGF-8 gene have been seen in patients with Cleft lip and/or cleft palate. This condition results from the mis-sense mutation D73H. It is thought that this mutation occurs at the N terminal of the FGF-8 protein and is important for the binding of FGF-8 to the receptor.
FGF-9 stimulates proliferation and activation of glial cells. FGF-9 has the most specificity to FGFR3 receptor; this makes it the most likely FGF to be involved in skeletal disorders. Six mutations have been found on FGF-9 that correlate to colorectal, endometrial, and ovarian cancers.
FGF-10 is the most closely related to FGF-7. Like FGF-7, FGF-10 is specific for epithelial cells. FGF-10 is involved in wound healing and embryonic development. Mutations in FGF-10 have been shown to cause autosomal dominant aplasia or lacrimal and salivary glands (ALSG) and lacrimo-auriculo-dento-digital syndrome (LADD).
FGFs 11-14 lack signal peptides but contain nuclear localization sequences. It appears that FGFs 11-14 play an important role in the nervous system. These FGFs are not activated by the FGF receptors. Instead, these FGFs mediate intracellular signaling.
It is believed that FGF-15 may play an important role in the nervous system. It has a 30% homology with the rest of the FGF family. Recently FGF-15 has been shown to play an important role in metabolism, specifically in bile acid synthesis. FGF-15 does not express its biological function in a paracrine manner. FGF-16 is most similarly related to FGF-9 by a homology of about 73%. It is thought to play a role in brown adipose tissue and the amount of FGF-16 in the body decreases significantly after birth. FGF-17 is most similarly related to FGF8. It is thought to be involved in embryonic brain development. This FGF isoforms is the most closely similar to FGF-8 and FGF-17 by about 53%.
Unlike other FGFs that play a role in proliferation, FGFs 19-23 play an important role in different types of metabolism. FGF-19 is used to regulate the metabolism in an endocrine manner and does not express its biological function in a paracrine manner. This protein regulates energy, lipid, and bile acid metabolism. It is thought that FGF-21 regulates energy, lipid, and glucose production. FGF-23 regulates phosphate and vitamin D metabolism. These FGFs can activate their receptors but are much less efficient and need the assistance of cofactors.
Structure of FGFs
FGFs contain more than one cysteine but these cysteines do not form disulfide bonds. All FGFs are a β-trefoil fold consisting of 12 antiparallel β strands arranged into three sets of four stranded β sheets. There are no a helical segments in the FGF structure. The length of β strand I vary significantly in the FGF family. Most of the β strands have similar amino acid composition throughout the FGF family; however, the loop regions vary throughout the FGF family. The loop region is responsible for most of the accessible surface of the FGFs. The amino acids that are not involved in the core of the β-trefoil structure of the protein on the far N and C terminal vary in amino acid sequence among the FGF family. These variations located on the N terminal and C-terminal ends make the biology of each FGF different.
Fibroblast Growth Factor Receptor (FGFR)
Many of the FGFs mediate their biological processes by binding to their receptor, FGFR. FGFRs are members of the tyrosine kinase family. There are currently five known isoforms of FGFR. The first four are similar in structure; each contains three immunoglobulin "Ig"-like domains D1-D3, a single transmembrane domain, and an intracellular tyrosine kinase domain. FGFR1-4 are high affinity, FGF ligand dependent proteins. Domains (D1 - D3) are the extracellular region of the receptor and are responsible for ligand binding specificity and receptor dimerization. These receptors are stabilized by heparin which then leads to the subsequent dimerization and activation of the FGF/FGFR complex. All four receptors are very similar in homology. FGFR1 is the largest of the four receptors; it contains a larger D1 domain and a longer D1-D2 linker. FGFR4 is the smallest receptor and contains the smallest D1 domain and the shortest D1-D2 linker. Each extracellular domain consists of two β sheets connected by a sulfide bridge. This extracellular region also contains a stretch of acidic amino acids between the D1 and D2 domain. This stretch of amino acids is known as "acid box".
Two splicing events occur in the FGF receptor, which gives the receptor specificity for each FGF. One splicing event involves the removal of D1 and or D1 along with the acid box region form the FGFR. The results of the remove of the first domain (D1), gives the shortened receptor a higher affinity of the FGF ligand. The other splicing event occurs in the D3 domain and involves mRNA splicing. This alternative mRNA splicing event gives different ligand binding properties to the receptor.
FGFR5 has 32% identity to the extracellular domain of FGFRs 1-4. Unlike other FGFR isoforms, FGFR5 only has two "Ig"-like domains. It also does not have an intracellular tyrosine kinase domain. FGFR5 can be found in variety of tissues including: kidney, brain, and lungs.
|FGFR Isoform||Ligand Specificity|
|FGFR1b||FGF1, 2, 3 and 10|
|FGFR1c||FGF1, 2, 4, 5 and 6|
|FGFR2b||FGF1, 3, 7, 10 and 22|
|FGFR2c||FGF1, 2, 4, 6, 9, 17 and 18|
|FGFR3b||FGF1 and 9|
|FGFR3c||FGF1, 2, 4, 8, 9, 17, 18 and 23|
|FGFR4||FGF1, 2, 4, 6, 8, 9, 16, 17, 18 and 19|
Fibroblast growth factor reference
1. Dailey L, Ambrosetti D, Mansukhani A, et al. Mechanisms underlying differential responses to FGF signaling[J]. Cytokine & growth factor reviews, 2005, 16(2): 233-247.
2. Itoh N, Ornitz D M. Functional evolutionary history of the mouse Fgf gene family[J]. Developmental Dynamics, 2008, 237(1): 18-27.
3. Baird A, Ornitz D M. Fibroblast growth factors and their receptors[J]. Angiogenesis in health and disease: basic mechanisms and clinical applications. New York: Marcel Dekker Inc, 2000: 75-88.
4. Zhu X, Komiya H, Chirino A, et al. Three-dimensional structures of acidic and basic fibroblast growth factors[J]. Science, 1991, 251(4989): 90-93.
5. Yeh B K, Igarashi M, Eliseenkova A V, et al. Structural basis by which alternative splicing confers specificity in fibroblast growth factor receptors[J]. Proceedings of the National Academy of Sciences, 2003, 100(5): 2266-2271.