Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

Filamin A

Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_101630

Synonyms

Historical Background

Filamin A was initially discovered in 1975 as a 280 kDa actin-binding protein in rabbit alveolar macrophages (Hartwig and Stossel 1975), and was named Actin-Binding Protein 280 (ABP280). In the same year, Wang and colleagues described a high-molecular-weight nonmuscle actin-binding protein in chicken gizzard extracts and called it filamin because it was localized on actin filaments (Wang et al. 1975). The suspicion that ABP280 and filamin were the same protein was confirmed 15 years later with the cloning of the ABP280/filamin gene (Gorlin et al. 1993). Today we know that mammals have three filamins; in addition to filamin A (gene symbol FLNA, chromosome Xq28), two other genes with high homology to filamin A were cloned in 1998 and 2000, respectively called filamin β and FLN2 (Takafuta et al. 1998, Thompson et al. 2000). FLNA is the most broadly expressed member of the filamin family but filamin β, subsequently renamed filamin B (gene symbol FLNB, chromosome 3p14), is also widely expressed while FLN2, renamed filamin C (gene symbol FLNC, chromosome 7q32), is largely restricted to skeletal and cardiac muscle. Although first appreciated primarily as actin-crosslinking proteins important for stabilizing branched actin networks, it is now recognized that filamins also play key roles as signaling scaffolds, as links between transmembrane receptors and the actin cytoskeleton, and in mechanosensing and mechanotransduction.

Filamin A Architecture

Filamins are composed of an N-terminal F-actin binding domain (ABD) followed by 24–100-residue immunoglobulin-like domains (IgFLN domains), the most C-terminal of which (IgFLN24) mediates homodimerization (Razinia et al. 2012) (Fig. 1). The filamin actin-binding domain is composed of tandem calponin homology domains (CH1 and CH2)) exhibiting sequence and structural similarities to the actin-binding domains of α-actinin, fimbrin utrophin, and distrophin (Razinia et al. 2012). The molecular basis for ABD interaction with actin is not yet fully understood, although conformational rearrangements between the CH1 and CH2 domains may be important to facilitate binding and prevent steric clashes. Notably, filamin A has an additional binding site located in IgFLN 10 (Nakamura et al. 2007). While the functional importance of this second actin-binding site is not completely clear, it seems to cooperate with the N-terminal ABD to allow maximum F-actin binding.
Filamin A, Fig. 1

Schematic representation of human filamin. The N-terminal actin-binding domain containing two calponin-homology domains (CH1 and CH2) is followed by 24 immunoglobulin-like repeats of ∼96 amino acids each. The repeats are interrupted by two hinge regions (hinge 1 and hinge 2). Repeats 1–15 are mostly arranged in a linear form and make up a rod-like structure (rod 1). Repeats 16–24 (rod 2) form pairs resulting in a more compact structure that is central to mechanical signaling by filamins. The C-terminal twenty-fourth repeat is the dimerization domain. The location of disease-associated missense mutations is indicated by color-coded dots. PH blue, OSD red, CVD green

Sequence analysis led to early predictions that filamin A was composed of repeating units (Gorlin et al. 1993), and structural studies on vertebrate and invertebrate filamins subsequently revealed that the repeating unit has an immunoglobulin-like fold composed of seven β-strands (A to G) assembled into a two β-sheet sandwich (Fig. 2). Notably, filamin A is reported to interact with more than 100 binding partners (Table 1), and the great majority of filamin interactions are mediated by IgFLN domains, particularly IgFLNa domains 16–24. Strikingly, the structures of IgFLN domains binding to peptide ligands reveal conserved modes of interaction with the ligand generally forming an additional antiparallel β-strand next to β-strand C of the CFG sheet (Kiema et al. 2006) (Fig. 2). In addition to characterizing IgFLN domain ligand interactions, structural studies have revealed how the tandem IgFLN domains are organized. The first fifteen IgFLN domains are thought to be arranged in a linear manner forming an elongated rod-like structure 58 nm in length (rod1, Fig. 1) while, in contrast, the 8 repeats from IgFLN16 to IgFLN24 (rod 2) form a more compact structure (19 nm in length) (Nakamura et al. 2007) (Fig. 1). Structural studies indicate that this compact structure is generated because IgFLNa 16 pairs with IgFLNa17, IgFLNa 18 pairs with IgFLNa19, and IgFLNa20 pairs with IgFLNa21 (Lad et al. 2007, Ruskamo et al. 2012). This organization is functionally relevant as pair formation can inhibit ligand binding to the odd-numbered IgFLN domain within the complex.
Filamin A, Fig. 2

Ribbon diagram of the crystal structure of IgFLN21 (green) interacting with a peptide from the cytoplasmic region of the β7 integrin (yellow) (PDB 2BRQ). Letters A–G indicate the β-strands of the filamin domain forming two sheets: GFC (on the front) and ABED (on the back). Note that the ligand forms an extra strand next to the C strand

Filamin A, Table 1

Filamin A binding partners

Interactor

Experimental system

Pubmed ID

 ITGA4

Affinity capture-MS

22623428

ITGB1

Two-hybrid; Reconstituted complex; Affinity capture-Western

11807098; 9722563

ITGB3

Two-hybrid

11807098

ITGB6

Two-hybrid

11807098

ITGB7

Reconstituted complex

11781567

KCND2

Affinity capture-western

20498229

KCNE4

Affinity capture-western; Two-hybrid

20498229

LGALS1

Cofractionation

22939629

LIG4

Affinity capture-MS

22990118

LMNA

Two-hybrid; Proximity label-MS

16248985; 22412018

MAPK14

Two-hybrid

20936779

MDC1

Affinity capture-MS

22990118

 MTNR1A

Affinity capture-MS

17215244

MYBL2

Affinity capture-MS; Cofractionation; Affinity capture-western

18548008

 MYC

Affinity capture-MS

21150319

MYH9

Cofractionation

22939629

MYOT

Affinity capture-western

11038172

NPHP1

Two-hybrid; Affinity capture-western

12006559

NPM1

Affinity capture-MS

23402259

OTUD1

Affinity capture-MS

19615732

OTUD5

Affinity capture-MS

19615732

PAN2

Affinity capture-MS

23398456

RALA

Affinity capture-western; Reconstituted complex

10051605

RANBP2

Cofractionation

22939629

RBM39

Cofractionation

22939629

RPS23

Cofractionation

22863883

 SH2B3

Affinity capture-western; Two-hybrid

11163396

SHBG

Two-hybrid

15862967

SIRT7

Affinity capture-MS

22586326

SMAD3

Two-hybrid

15231748

SMURF1

Affinity capture-MS

23184937

 SRC

Two-hybrid

20936779

SRRM2

Cofractionation

22939629

SRSF11

Cofractionation

22939629

STK4

Affinity capture-MS

23455922

SUMO1

Reconstituted complex

19596686

SUMO2

Affinity capture-MS

21693764; 19471022; 21252943; 23077236;

SVIL

Two-hybrid

20309963

SYNPO2

Affinity capture-western

23434281

TJP1

Affinity capture-MS

16944923

TP53BP1

Affinity Capture-MS

22990118

TRA2B

Cofractionation

22939629

TRAF2

Affinity capture-western

10617615

 TRIO

Reconstituted complex

11146652

TSGA10

Affinity capture-MS

20797700

UBC

Affinity capture-MS

16335966; 18781797; 21987572; 21139048; 21890473; 21906983; 22505724; 21963094; 22053931; 23000965

UBD

Affinity capture-MS

23862649

USP19

Two-hybrid

23500468

USP45

Affinity capture-MS

19615732

VCAM1

Affinity capture-MS

19738201

VHL

Two-hybrid

8674032; 12169691; 11146652

Filamins are classically described as having two flexible hinge regions, the first between repeats 15 and 16 separating rod 1 and rod 2 and the second preceding IgFLN24 (Fig. 1), although alternative splicing can influence the presence of hinge regions in filamin B. Calpain proteolysis in the first hinge region generates fragments of approximately 190 kDa (ABD to domain 15) and 90 kDa (domain 16–24). The 90 KDa fragment resulting from calpain cleavage is reported to localize in the nucleus and interact with nuclear receptors (McGrath et al. 2013). The physiological relevance of calpain cleavage in the second hinge region is not clear.

Filamin A and Mechanotransduction

It is now recognized that mechanical cues are as important as chemical signals in controlling cell-fate decisions and spatiotemporal regulation of transcription. Cell surface receptors convey extracellular mechanical signals to the cytoskeleton allowing the actin cytoskeleton to rapidly transfer them through the cell, and a range of studies implicate filamin in this process (Razinia et al. 2012). Notably, the recent studies of mechanically strained filamin–crosslinked actin networks (Ehrlicher et al. 2011) and single molecule measurements (Rognoni et al. 2014) support a force-sensing role for filamin and together with structural and functional studies (Lad et al. 2007) have shed light on potential molecular mechanisms. As noted above, Filamin A interacts with more than 100 partners (Table 1) to affect a wide range of cellular activities, with the majority of interactions occurring in rod 2. In the domain pairs found in rod 2, the A-strand of the even-numbered domain can bind to the ligand-binding site of the subsequent odd-numbered domain inhibiting the interaction of this domain with its ligands (Fig. 3). When force is applied along the filamin A molecule, the conformational equilibrium of the autoinhibited filamin domain pairs is shifted to displace and eventually unfold the inhibitory strand from the adjacent domain with a subsequent increase in affinity of that domain for ligands (Fig. 3) (Ehrlicher et al. 2011, Rognoni et al. 2014).
Filamin A, Fig. 3

Model for the effect of mechanical stress on filamin rod 2. Force will lead to domain pair opening allowing binding. Under high stress FLNa20 unfolds, exposing ligand-binding sites hidden in low stress conditions

Filamin A in Human Disease

Mutations in each of the three human filamin genes have been associated with a wide spectrum of congenital anomalies including malformations of the skeleton, brain, muscle, and cardiovascular system (Nakamura et al. 2011; Robertson 2005). This indicates that, despite the conservation between filamins and their often-overlapping expression patterns, each filamin has a specific function that cannot be fully compensated by the other isoforms. In particular, mutations in the filamin A gene cause a wide range of human disease symptoms. This broad phenotypic presentation is likely to be explained by the observation that mutations on the filamin A gene can exert both gain- and loss-of-function effects on the protein. In addition, the location of FLNA on the X chromosome explains differences in phenotypic presentation between genders as well as variations in symptom severity due to random X-chromosome inactivation.

The first disease to be associated with FLNA mutations was Periventricular nodular heterotopia (PH). PH is an X-linked disease characterized by incomplete radial migration of cortical neurons and is described in females who are heterozygous for loss-of-function FLNA mutations. The defective neuronal migration is often associated with seizures but can also cause cognitive disorders. In addition, PH is associated with vascular anomalies such as patent ductus arteriosus and cardiac valvular anomalies, and is often accompanied by megathrombocytopenia and joint hypermobility. The elevated number of early miscarriage and an underrepresentation of males within PH kindreds suggest an early embryonic lethality in males who are hemizygous for a truncating FLNA mutation. The few cases of PH in males report more severe phenotypes both at neuronal and cardiovascular level. For a decade, PH was considered to reflect a general migration defect in filamin A deficient cells. However, this hypothesis has been subsequently challenged by studies revealing normal migration in filamin A knockdown or knockout cells in vitro and normal neuronal migration in mice (Baldassarre et al. 2009). This may be due to compensation by other filamin isoforms because, while loss of one single isoform (filamin A or filamin B) has no effect on cell migration in many cell types, knockdown of both results in impaired initiation of migration and spreading (Baldassarre et al. 2009). Why loss of filamin A causes PH in humans remains unclear.

Five years after the discovery that filamin mutations cause PH, filamin A mutations were also linked to Otopalatodigital Spectrum Disorders (OSD, otopalatodigital syndrome-1, otopalatodigital syndrome-2, Frontometaphyseal dysplasia, and Melnick-Needles syndrome) (Robertson et al. 2005). OSD are characterized primarily by a skeletal dysplasia of variable severity in conjunction with malformations in other organ systems including the heart, respiratory tract, and gastrointestinal system. As seen for PH, females typically have milder OSD symptoms, mainly affecting the skeleton. In contrast, males have more severe skeletal defects with frequently life-limiting extraskeletal malformations such as cleft palate, septal heart defects, omphalocoele, and renal tract malformations (Robertson 2005). Mutations linked to these diseases are usually missense point mutations that are hypothesized to confer gain of function to the protein. As shown in Fig. 1, mutations associated with OSD cluster in specific “hot spots” on the protein suggesting alteration of specific functions of the protein causes disease. The concentration of mutations in the ABD and domain 10 (Fig. 1) along with detailed studies of one OSD mutant filamin (Clark et al. 2009) suggests that increased filamin A binding to F-actin may be responsible for the development of OSD phenotypes. Mechanisms by which increased actin binding would cause OSD remain to be determined.

Recently filamin mutations have also been linked to X-Linked Chronic Idiopathic Intestinal Pseudo-obstruction (CIPO), a condition described mainly in males with severe intestinal dysmotility often accompanied by perinatal mortality and shortening of the small intestine. Mutations linked to CIPO are likely to lead to reduced expression of FLNA although gene duplication has also been reported (Clayton-Smith et al. 2009).

Cardiac valvular dysplasia, x-linked (CVD), a rare form of cardiac valvular dystrophy that is not associated with PH or OSD is also linked to missense filamin A mutations or a large inframe deletion. As with other filamin A-associated diseases, phenotypes are more severe in males where all the valves can be affected while heterozygous female are considered asymptomatic with only very mild phenotypes (Kyndt et al. 2007).

In addition to the abovementioned genetic diseases, alterations in filamin A levels have been linked to cancer progression and dissemination. Although filamin A was initially considered a proinvasion protein, more recent studies have shown that its expression negatively correlates with metastatic potential in breast cancer (Xu et al. 2010). In vitro filamin A depletion decreases secretion of the tissue inhibitor of metalloproteinase and consequently increases activation of matrix metalloproteinase 2 and cell invasion (Baldassarre et al. 2012), potentially explaining the effects on metastasis in vivo. Furthermore, filamin A is reported to inhibit the transcriptional activity of the androgen receptor and decrease the growth of prostate cancer (McGrath et al. 2013).

Summary

Filamin A is an important actin-binding protein with essential roles in development, tissue formation, and morphogenesis. Consistent with this, mutations in filamin A have been associated with a wide spectrum of congenital anomalies including malformations of the skeleton, brain, muscle, and cardiovascular system, and filamin A plays important and complex roles in cancer. Filamin A functions as a noncovalent homodimer of 240–280 kDa subunits composed of an N-terminal actin-binding domain followed by 24 immunoglobulin-like (IgFLN) domains, the last of which mediates dimerization. A wide range of biochemical, biophysical, structural, and cell biological studies have provided significant insights into the mechanisms by which filamins regulate cell behavior and mediate mechanosensing, and more than 100 filamin-binding partners have been reported. However, the physiological significance of many reported interactions remains unclear and exactly how filamin mutations cause human disease is an important topic for future study.

References

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Copyright information

© Springer International Publishing AG 2018

Authors and Affiliations

  • Massimiliano Baldassarre
    • 1
  • David A. Calderwood
    • 2
  1. 1.University of Aberdeen Institute of Medical SciencesAberdeenUK
  2. 2.Departments of Pharmacology and of Cell BiologyYale School of MedicineNew HavenUSA