Intrinsic Disorder in BAP1 and Its Association with Uveal Melanoma

Abstract

Background: Specific subvariants of uveal melanoma (UM) are associated with increased rates of metastasis compared to other subvariants. BRCA1 (BReast CAncer gene 1)-associated protein-1 (BAP1) is encoded by a gene that has been linked to aggressive behavior in UM. Methods: We evaluated BAP1 for the presence of intrinsically disordered protein regions (IDPRs) and its protein−protein interactions (PPI). We evaluated specific sequence-based features of the BAP1 protein using a set of bioinformatic databases, predictors, and algorithms. Results: We show that BAP1’s structure contains extensive IDPRs as it is highly enriched in proline residues (the most disordered amino acid; p-value < 0.05), the average percent of predicted disordered residues (PPDR) was 57.34%, and contains 9 disorder-based binding sites (ie. molecular recognition features (MoRFs)). BAP1’s intrinsic disorder allows it to engage in a complex PPI network with at least 49 partners (p-value < 1.0 × 10−16). Conclusion: These findings show that BAP1 contains IDPRs and an intricate PPI network. Mutations in UM that are associated with the BAP1 gene may alter the function of the IDPRs embedded into its structure. These findings develop the understanding of UM and may provide a target for potential novel therapies to treat this aggressive neoplasm.

Keywords: BAP1; intrinsically disordered protein (IDP); intrinsically disordered protein regions (IDPR); protein–protein interaction network; uveal melanoma.

Figure 1

Structures generated for BAP1 by AlphaFold2. Regions with higher model confidence as measured by the predicted local distance difference test (pLDDT) typically represent α-helices. Regions with lower model levels represent intrinsically disordered protein regions. The black arrows indicate areas of high levels of disorder and lack of structure.

Figure 2

Amino acid composition profile of BAP1. The fractional difference is calculated as (Cx–Corder)/Corder, where Cx is the content of a given amino acid in the query set (BAP1), and Corder is the content of a given amino acid in the background set (Protein Databank Select 25). The amino acid residues are ranked from most order promoting residues ((i.e., cysteine (C), tryptophan (W), isoleucine (I), tyrosine (Y), phenylalanine (F), leucine (L), histidine (H), valine (V), asparagine (N), and methionine (M)) to most disorder promoting residues ((i.e., arginine (R), threonine (T), aspartate (D), glycine (G), alanine (A), lysine (K), glutamine (Q), serine (S), glutamate (E), and proline (P)). Positive values indicate enrichment, and negative values indicate depletion of amino acids. 7 of 10 order-promoting residues are depleted and 5 of 10 disorder-promoting residues are enriched. Residue bars marked with * are statistically significant for depletion (i.e., W) or enrichment (i.e., S and P) (p-value < 0.05)).

Figure 3

Evaluation of the intrinsic disorder predisposition of human BAP1. (A) Per-residue intrinsic disorder profiles generated by six commonly used intrinsic disorder predictors, PONDR^® VLXT, PONDR^® VL3, PONDR^® VSL2, PONDR^® FIT, IUPred short, and IUPred long. The thin black line at the disorder score of 0.5 is the threshold between order and disorder, where values above 0.5 are disordered residues and values below 0.5 are ordered residues. Thin black dashed line at disorder score 0.15 shows the threshold between flexible and mostly ordered region/residues. There are many segments of disorder seen throughout BAP1’s structure. In particular, the amino acid segment from 250 to 500 is highly enriched with intrinsic disorder. (B) The D2P2 platform output is used to assess the functional disorder profile for BAP1 protein. On the left of the figure, there are four identifiers for predictors used on the D2P2 platform. The top segment shows outputs from various per-residue disorder predictors, the second segment shows predicted protein domains, the third segment shows where molecular recognition features are located (MoRFs; i.e., disorder regions that become ordered when binding), and the fourth segment shows posttranslational modifications (PTM) sites. The per-residue predictors agree and align with the PONDR^® outputs from (A). The protein domains fall within regions that are predicted to be ordered or disordered. The MoRF regions are localized to regions of that demonstrate extensive disorder. The PTMs are also predominantly localized to areas of BAP1 that contain intrinsic disorder.

Figure 4

Search Tool for the Retrieval of Interacting Genes (STRING) output for BAP1. The graphic demonstrates that BAP1 (black circle) interacts with (lines) multiple proteins (colored circles). The ability for BAP1 to interact with 49 other proteins is likely possible through the utilization of IDPRs. The number of edges in this protein interaction network is 599, and the expected number of edges for randomly selected proteins of the same-sized network is 101 (p-value < 1.0 × 10⁻¹⁶).