Intrinsic Disorder and the Evolution of Overlapping Genes

THE TEAM: Pedro R. Romero1, Corinne Rancurel2, Mahvash Khosravi1, Keith Dunker1, and David Karlin3
2Architecture

University - Purdue University Indianapolis, Indianapolis, IN, USA, et Fonction des Macromolcules Biologiques, Campus de Luminy, Marseille, France 3Tous Chercheurs, Inmed, Parc de Luminy, 13273 Marseille Cedex 09, France

1Indiana

Overlapping Genes and Overprinting

Overlapping genes discovered in first sequenced genome (Phage -X174, Sanger, 1977) Many theoretical studies in the 70s
Constrained evolution demonstrated

Limited information content/gene (Yockey, 78)

Predicted difference in evolutionary speed (Smith & Waterman, 78)

Overprinting of older genes proposed as mechanism for de novo gene generation (Keese & Gibbs, 1992) Structural effects noticed in isolated examples: some recent ones shown to be related to intrinsic disorder.

Overprinting: Creation of a novel C-terminal extension

Overprinting Examples

Overprinting and Structure Example: Measles virus (Karlin, et al, 2002)

view of the measles virus. The nucleocapsid protein (N) assembles into a wraps the virus RNA. The nucleocapsid protein has a disordered tail where ) binds. The P protein is largely disordered, and it is encoded by a multipleRNA (see Figure 2).

Figure 2. Schematic view of the RNA region that encodes the P, V, and C proteins in the measles virus genome. The color lines at the bottom of the diagram represent the three RNA reading frames. The phosphoprotein (P) and the N-terminus of the V protein are encoded on frame 1 (blue), the Cterminus of the V protein is encoded on frame 2 (red), and the entire C protein is encoded on frame 3 (green). The colored cylinders at the top represent the encoded protein products, with the narrow cylinders denoting disordered regions and the wide cylinders representing ordered regions. Notice that no region in the RNA encodes two ordered protein domains.

Intrinsically Disordered Proteins

IDPs and regions are less sensitive to evolutionary changes than ordered ones They undergo faster evolution than ordered proteins or regions More dependent on residue content than position Mostly participants in regulatory and signaling functions

Structural analysis of viral overlapping genes

Hypotheses 1. Intrinsic disorder might help alleviate evolutionary constraints in overlapping genes 2. New, overprinted genes will likely tend to be more disordered than ancestral counterpart

Structural analysis of viral overlapping genes

Need for real, expressed genes Spliced genes excluded (many unannotated splicing events) Assembled data from 43 viral genomes
Unspliced RNA virus Unspliced retroid virus (RNA and DNA) Overlaps > 90 nucleotides (30 residues)

Analysis of viral overlapping genes

Analysis of viral overlapping genes

Table 2. Predicted order/disorder statistics on overlapping genes data set.

Measures of order content Fraction of sequence predicted ordered Entire data set Encoded by overlapping genes Fraction of overlapping sequence positions predicted ordered on both protein products Expected Observed

Fraction 71% 52%

Confidence Intervals 68% (1 std. dev.) 95% 68-74% 48-57% 66-76% 45-60%

50% 28%
0.6

46-55% 23-33%

44-58% 19-36%

FractionO-O

Difference in disorder content between entire data set and overlapping regions is significant (p-value 3x10-57) Difference between expected and observed fraction of order-order overlaps is also significant (P-value 5x10-24)

0.5

0.4

0.3

0.2

0.1

0 Expected Observed

Structural and functional organization of ancestral/novel proteins

Structural and functional organization of ancestral/novel proteins

Structural and functional organization of ancestral/novel proteins

Overprinting (novel) proteins
Most are Orphans (no homologs outside of genus) Mostly disordered Mostly accessory proteins Proteins created by overprinting different homologs of the same gene display a wide diversity of functional and of structural features

Disorder and evolutionary constraints

100%

More constrained Less constrained

% disorder in overlapping region

80%

60%

40%

20%

0%
HBV Sendai SIV HTLV phiX174 phiX174 A-B D-E PLRV HPV CLCuV AC1AC4 CLCuV CP-AV2

Virus

Disorder and evolutionary constraints

100%

More constrained Less constrained

% disorder in overlapping region

80%

60%

40%

20%

0%
HBV Sendai SIV HTLV phiX174 phiX174 A-B D-E PLRV HPV CLCuV AC1AC4 CLCuV CP-AV2

Virus

Disorder and evolutionary constraints

Experiment shows that less constrained (faster evolving) proteins in a pair tend to be more disordered Only exceptions when disorder content very similar between the two overlapping proteins

Conclusions
Both proposed hypothesis supported:
Disorder appears to alleviate evolutionary constraints Novel overprinted genes tend to be disordered

New directions: Show whether novel genes tend to be disordered at birth or are selected to be disordered

THANKS!!

ematic view of the measles virus. The nucleocapsid protein (N) assembles into a id that wraps the virus RNA. The nucleocapsid protein has a disordered tail where otein (P) binds. The P protein is largely disordered, and it is encoded by a multiple-