Fig 1.
The four components of our bioinformatics strategy.
(A) Identify human-type short linear motifs (SLiMs) found in a viral protein(s). (B) Construct a virus-host-PPI network. (C) Identify network modules and roles of the network nodes. (D) Build a map of AVP/SLiM-protein complexes. In parentheses are the numbers of motifs, proteins, modules, pathways, etc., identified in this study (see Methods for details).
Fig 2.
Modules and roles of the nodes of the HCV-Human PPI network.
The network was produced using the procedures described in Fig 1A–1C, with the modules and the roles of network nodes determined by NetCarto (see Methods). The three types of network nodes are represented as triangles for SLiMs; circles for VIPsdirect; and squares for VIPsindirect. Their network roles (R7 to R1) are depicted as symbols of decreasing size. All nodes are color-coded according to their module. A representative function(s) of each module was derived from an enrichment analysis of GO terms associated with its nodes followed by a summary of Revigo representatives [21] (see Methods and S1 Table). Colored in gray in the lower right corner are eight isolated nodes.
Fig 3.
Relationships between SLiMs, R6 VIPs, and network modules.
Of the 19 SLiMs identified for HCV E1 and E2 in Fig 1C, 13 (including six grouped in the MOD_family; top row) are directly connected to one or more of the 22 R6 VIPsdirect (middle row) in the virus-host PPI network (Fig 2). The MOD_family contains MOD_CK1_1, MOD_CK2_1, MOD_GSK3_1, MOD_NEK2_1, MOD_NEK2_2, and MOD_ProDKin_1; all are targets of a kinase. An R6 VIPdirect and a module(s) (bottom, boxed in black) are considered to be connected if more than 10% of the interacting partners of the VIPdirect belong to the module. Based on this criterion, module 2 is not connected to an R6 VIPdirect and, therefore, is not included in the figure. The validity of the connections displayed as solid dark lines is supported by published experimental evidence. The corresponding reference number(s) (indicated by an asterisk) are provided in S3 Table. The dark and light horizontal bars at the bottom of the figure identify modules with the functionality of entry and/or carcinogenesis, respectively.
Table 1.
Comparison with known entry factors of HCV infection.
Fig 4.
The AVP/SLiM-protein complex map.
Top panel: (left column) a list of AVPs with their AVPdb identification numbers that have an amino acid sequence containing one or more HCV E1/E2 SLiMs (indicated by the triangles and named at the bottom of the panel) that can bind to a subunit of a protein complex belonging to one of the six main complex groups. The relative efficacies of these AVPs in inhibiting HCV entry according to data provided in the AVPdb [56] are indicated by the circles column in the panel with the shading score shown to the right of the panel. Middle panel: the network connecting the nine SLiMs to their group A-F complex target(s). Within each group are VIPs of all the complexes: ovals represent VIPsdirect (gray ovals are R6 VIPsdirect), and rectangles represent VIPsindirect (not all VIPsindirect are shown). The ovals and rectangles in bold outline are known HCV entry factors. The horizontal bar represents the plasma membrane, the VIPs below the bar and within the shaded area are ‘peripheral membrane proteins’, and those spanning the bar are ‘integral membrane proteins’. APOE is a peripheral membrane protein located at the extracellular side of the cell membrane. A connection between a viral SLiM and a complex group indicates that at least one protein in the group is targeted by HCV E1 and/or E2 via the viral SLiM. The thickness of the connection roughly scales to the number of proteins targeted by the SLiM in the group. The three numbers in the parenthesis are the number of VIPsdirect, total VIPs (i.e., VIPsdirect plus VIPsindirect), and unique subunits in the complex group. Bottom panel: the KEGG pathways enriched in the complex group (Benjamini-Hochberg adjusted P < 0.001) (see S6 Table for the pathway names). The gray scale at bottom right indicate the significance of the P-values. The functions of the individual KEGG pathways are shown at the left of the panel and the main functionality at the right of the panel.
Fig 5.
(A) The EGFR-associated complex (S5 Table, Complex ID: 176). (B) The AKT1-associated complex (S5 Table, Complex ID: 180). (C) Types of SLiM-domain interactions (indicated by (1), (2), and (3)) that mediate the targeting of protein complexes, and three sets of AVPs (indicated by (I), (II) and (III)) containing the corresponding SLiM that exhibits inhibition activity (indicated by bar-headed arrows) are shown. Proteins represented by black ovals are VIPsdirect; by gray ovals are VIPsindirect; and by white ovals are not VIPs (i.e. not in the virus-host PPI network). Double-headed arrows in panel A and B indicate predicted HCV-host PPIs in our analysis. Viral SLiM(s) are highlighted within the amino acid sequence of the AVP and are accompanied by its relative efficacy (in shaded circles) in inhibiting HCV entry (see the vertical bar for the normalized inhibition score in Fig 4 on the right of its top panel). The regular expression of each SLiM sequence as annotated in the ELM database [11] is shown within quotation marks in the SLiM-domain interactions (1), (2), and (3). *EGFR is a known HCV entry factor. **MOD_ProDKin_1 is representative of the MOD_family SLiMs (see Fig 3).
Fig 6.
Definitions and schematic depictions of network roles.
According to NetCarto [18], nodes (small red circles) with z-score of within-module degree ≥ 2.5 are defined as module hubs (nodes with many links, i.e. spikes in the schematic view), and those with z-score < 2.5 are non-hubs. Large circles represent modules. See [18] for further details.