Negative selection maintains transcription factor binding motifs in human cancer

Abstract

Background: Somatic mutations in cancer cells affect various genomic elements disrupting important cell functions. In particular, mutations in DNA binding sites recognized by transcription factors can alter regulator binding affinities and, consequently, expression of target genes. A number of promoter mutations have been linked with an increased risk of cancer. Cancer somatic mutations in binding sites of selected transcription factors have been found under positive selection. However, action and significance of negative selection in non-coding regions remain controversial.

Results: Here we present analysis of transcription factor binding motifs co-localized with non-coding variants. To avoid statistical bias we account for mutation signatures of different cancer types. For many transcription factors, including multiple members of FOX, HOX, and NR families, we show that human cancers accumulate fewer mutations than expected by chance that increase or decrease affinity of predicted binding sites. Such stability of binding motifs is even more exhibited in DNase accessible regions.

Conclusions: Our data demonstrate negative selection against binding sites alterations and suggest that such selection pressure protects cancer cells from rewiring of regulatory circuits. Further analysis of transcription factors with conserved binding motifs can reveal cell regulatory pathways crucial for the survivability of various human cancers.

Keywords: Cancer somatic mutations; DNA motifs; Negative selection; Transcription factor binding sites.

Fig. 1

Procedure used to evaluate affinity change events and estimate significance of difference between observed and expected frequencies. Top panel: prediction of binding sites in cancer and control data and evaluation of affinity change events. Bottom panel: binding sites predictions and affinity change events of AP-2 motifs; an example of 2 × 2 contingency table used to compute Fisher’s exact test P-value

Fig. 2

Selection magnitude for affinity loss and gain of ETS, FOX and C/EBP motifs in different cancer types. X-axis displays the selection magnitude for motif affinity loss (a) or gain (b) caused by somatic mutations. Box-plots are provided for ETS-related (14 motifs), FOX (13 motifs), C/EBP-related (9 motifs) and NR3 (Steroid hormone receptors, 11 motifs) transcription factor families in three cancer types with the largest numbers of mutation calls. In particular, C/EBP motifs display frequent affinity loss in breast cancer, FOX and NR3 motifs are protected from both the affinity loss and gain in lung adenocarcinoma and breast cancer, and ETS motifs tend to emerge in all three cancer types (breast, lung and liver). Data for two control datasets (shuffle, genomic) are shown

Fig. 3

Transcription factor binding motifs protected from somatic mutations in different cancer types. The size of a pie shows the total number of motifs in a given transcription factor family (given in curly braces according to TFClass). The slices of a pie show the number of conserved binding motifs protected from any affinity change (yellow), motifs protected from affinity loss (magenta), and motifs protected from affinity gain (deep purple)

Fig. 4

Procedure used to evaluate mutation frequencies relative to motif occurrences

Fig. 5

Relative location of mutated bases in reference to the AP2A (top panel) and ESR1 (bottom panel) binding motifs predicted for breast cancer data. Y axis shows the relative fraction of mutation-centered windows with the legitimate motif predictions, X axis shows the location of a mutated base relative to the motif. Motif logos are scaled according to the discrete information content. Somatic variants tend to localize at strict position of the AP2A motif (red line) leading to affinity change. Variants in ESR1 motif (purple line) avoid strict and prefer degenerate positions, the motif is protected from affinity change

Fig. 6

Fold change (log2) of negative selection magnitude for mutations in DNase accessible subregions compared to that in the promoter and intronic segments. Y axis displays selection magnitude fold change (log2), the ratio between selection magnitudes estimated for DNase accessible regions to those for all promoter and intronic segments, the respective genomic control data is used in the both cases. Lower values of selection magnitude correspond to the stronger negative selection, thus negative fold change values correspond to stronger negative selection in DNase accessible regions. X axis displays different significantly conserved motifs (P < 0.05) for the set of promoter and intronic mutations. Data for affinity loss (a, top panel) and affinity gain (b, bottom panel) is presented for breast cancer (top subpanels) and lung adenocarcinoma (bottom subpanels). Members of FOX and NR transcription factor families are colored in blue and green