The TRIOBP Isoforms and Their Distinct Roles in Actin Stabilization, Deafness, Mental Illness, and Cancer

Abstract

The TRIOBP (TRIO and F-actin Binding Protein) gene encodes multiple proteins, which together play crucial roles in modulating the assembly of the actin cytoskeleton. Splicing of the TRIOBP gene is complex, with the two most studied TRIOBP protein isoforms sharing no overlapping amino acid sequence with each other. TRIOBP-1 (also known as TARA or TAP68) is a mainly structured protein that is ubiquitously expressed and binds to F-actin, preventing its depolymerization. It has been shown to be important for many processes including in the cell cycle, adhesion junctions, and neuronal differentiation. TRIOBP-1 has been implicated in schizophrenia through the formation of protein aggregates in the brain. In contrast, TRIOBP-4 is an entirely disordered protein with a highly specialized expression pattern. It is known to be crucial for the bundling of actin in the stereocilia of the inner ear, with mutations in it causing severe or profound hearing loss. Both of these isoforms are implicated in cancer. Additional longer isoforms of TRIOBP exist, which overlap with both TRIOBP-1 and 4. These appear to participate in the functions of both shorter isoforms, while also possessing unique functions in the inner ear. In this review, the structures and functions of all of these isoforms are discussed, with a view to understanding how they operate, both alone and in combination, to modulate actin and their consequences for human illness.

Keywords: TRIOBP; actin; cancer; cytoskeleton; deafness; disordered structure; hearing loss; mental illness; protein aggregation; schizophrenia.

Figure 1

(a) Scale schematic of the alternative splicing of TRIOBP in humans. Exons (vertical bars) on the four most studied isoforms are shown, with introns represented by horizontal lines. Blue exons are entirely or mainly coding, black exons are entirely or mainly non-coding. Exon numbering is according to Park et al. [6]. (b) Scale schematic of the human TRIOBP-1, 4, 5, and 6 proteins with structural regions highlighted: R1, R2: First and second repeat domains, PH: Pleckstrin homology domain, Central: Central coiled coil domain, CT: C-terminal coiled coil domain. The number of amino acids (AA) in humans is also indicated. (c) The level of conservation of each section of the TRIOBP-6 amongst mammalian orthologues and predictions of three forms of secondary structure: disordered/unstructured protein, α-helix, and β-sheet. These are displayed as heat maps to scale with the schematic in part (b). Conservation determined using AL2CO [7], based on amino acid sequences of TRIOBP-6 (or similar splice variants) from 57 different mammalian genera. These were identified using BLAST (reference sequence human: TRIOBP-6, NP_001034230.1), aligned with CLUSTAL Omega 1.2.4 [8] and the alignment was then manually curated. Secondary structure predictions were made using PSIPDRED 4.0 and DISOPRED3 [9,10,11] with protein analyzed in three overlapping sections. All results were averaged over an 11 amino acid sliding window for clarity. The N-terminal 61 amino acids of TRIOBP-1 from exon 11a that are not present in TRIOBP-6 were not evaluated here, but were previously predicted to be disordered with comparatively poor conservation [12].

Figure 2

A structural homology model of the PH domain of TRIOBP-1 (amino acids 64–172 of 652 amino acid TRIOBP-1). (a) Images of the model, with the first loop region displayed. (b) Amino acid sequence of the PH domain, with the first and second loop regions indicated. Coloring corresponds to the secondary structures seen in the molecular images. (c) Image of the model including a low quality prediction of the strength of the second loop region. Model generated using MODELLER 9.20 [16], based principally on the structure of the PH domain of DAPP1 (PDB ID: 1FAO), which includes sequence analogous to the first loop region. Shorter sections including the second loop were modeled with additional templates (PDB ID: 2DYN, 2D9Y, 3GOC, and 5YUG). Alignments were generated using CLUSTAL Omega 1.2.4 [8], and then optimized manually. Of the 20 models generated, the one with the lowest objective function score was visualized using YASARA 18.4.24 [17].

Figure 3

The structure of the coiled-coil regions of TRIOBP-1. All parts of this figure are to scale with each other. (a) Locations of predicted coiled-coils (CC). Solid filled boxes represent high confidence predictions, striped boxes represent lower confidence predictions, derived from PSIRPED [9]. CCs are colored based on their predicted inclusion in the central CC domain (blue) or C-terminal CC domain (red). Amino acid numbering from both the 593 amino acid and 652 amino acid TRIOBP-1 proteins are shown. Labeling of CCs is based on Bradshaw et al. [12] and differs from the numbering used by Katsuno et al. [18], who do not count the putative coiled-coil labeled here as CC1 in their numbering. Level of amino acid conservation is displayed using the same calculation and heat map as in Figure 1c. (b) Locations of constructs representing the central and C-terminal CC domains from two publications [12,14]. (c) Locations of regions of TRIOBP-1 involved in protein–protein interactions and functions [3,12,14,19,20]. Note that some proteins bind more than one region of TRIOBP-1. The only proteins so far reported to bind to TRIOBP-1 outside of these CC regions is TRIO, which binds to the mid domain between the central CC region and the PH domain [13]. The locations of two known phosphorylated residues and their associated kinases are also shown [14,21]. (d) Locations of fragments of TRIOBP-1 and the oligomeric states they adopt when expressed in isolation in vitro [12].

Figure 4

(a) The location of the repeats that make up the R1 and R2 regions of TRIOBP-4, with ¸amino acid numbering of both TRIOBP-4/5 and TRIOBP-6. (b) The location of frameshift and nonsense mutations from patients with hearing loss. Red bars indicate homozygous mutations, while purple bars joined by dotted lines indicate compound heterozygous mutations. Arrowheads indicate that the other heterozygous mutations lie in a region of TRIOBP-5/6 that is 3′ of the TRIOBP-4 open reading frame. Full details of these are in Table 1. All elements of this figure are shown to scale.

Figure 5

Illustrated representation of the expression and roles of TRIOBP-1 (green), TRIOBP-4 (blue), and TRIOBP-5/6 (red). Clockwise from top: Aggregation of TRIOBP-1 in the brain and mental illness; Role of TRIOBP-1 in F-actin stabilization throughout the body; Linking of actin to ion channel function in the heart by TRIOBP-1; role of TRIOBP-1 in the cell cycle; Importance of all major TRIOBP isoforms in metastasis; Distinct roles of TRIOBP-4 and TRIOBP-5 in the stereocilia of the inner ear and deafness. Protein interaction partners implicated in the various processes are indicated in gray.