Systematic analysis of the frequently amplified 2p15-p16.1 locus reveals PAPOLG as a potential proto-oncogene in follicular and transformed follicular lymphoma

Abstract

Transformed follicular lymphoma (tFL) originates from histological transformation of follicular lymphoma (FL), which is the most common indolent non-Hodgkin lymphoma. High-resolution genomic copy-number analysis previously identified frequent amplification of the 2p15-p16.1 locus in FL and tFL cases. The genes (i.e. BCL11A, PAPOLG, PUS10, and USP34) in this amplified locus have not been systematically investigated to date in terms of their role in FL pathogenesis or transformation to tFL. Here we investigated the relationship between amplification and expression of genes in 2p15-p16.1 as well as their expression after histological transformation. NCBI GEO SNP array and gene expression profile (GEP) data of tFL cases were analyzed to evaluate the relationship between amplification and mRNA expression. Moreover, transcript levels of these four genes in FL cases were compared with those of patient-matched tFL cases and normal B-cells. Amplification of the 2p15-p16.1 locus is associated with increased transcription of BCL11A and PAPOLG in tFL cases, of which the latter showed increased expression after histological transformation. Compared with the level in normal B-cells, PAPOLG was significantly overexpressed in FL cases, but expression levels of the other three genes did not show any significant difference. Altogether these results suggest that PAPOLG may be the most critical gene in terms of transformation to tFL.

Keywords: 1; 2p15-16; Amplification; BCL11A; PAPOLG; PUS10; USP34; proto-oncogene; tFL.

Figure 1

Copy number estimates of four genes located in the 2p15-p16.1 locus in transformed follicular lymphoma cases. Gene copy numbers were estimated by calculating the average SNP array (Affymetrix mapping 250K Nsp SNP array) probe values for BCL11A (A), PAPOLG (B), PUS10 (C), and USP34 (D) for each of 42 tFL cases for which corresponding DNA microarray data are also available. Means ± SD for values of SNP array probes are shown for each gene. SNP probes used to estimate the copy number of each gene are described in the Materials and methods section. SD: Standard deviation.

Figure 2

Schematic depiction of the strategies used for mRNA expression analyses of amplified genes in follicular or transformed follicular lymphoma cases. A) Transformed follicular lymphoma cases were divided into two groups based on the high or low copy number of BCL11A, PAPOLG, PUS10, and USP34, which is determined using the values of normalized SNP array data. mRNA expression values were then analyzed with the GEO2R bioinformatics tool using the available DNA microarray probe sets (GEO accession number: GSE81184); B) Transcript expression of four amplified genes was evaluated with the GEO2R tool by comparing the expression levels in FL cases vs. normal B cell subsets (GEO accession number: GSE55267); C) Transcript expression levels of the four amplified genes in 12 tFL cases (i.e. DLBCL) were compared one by one with those of pretransformation levels for each tFL case by using the GEO2R tool (GEO accession number: GSE3458).

Figure 3

Amplification of BCL11A or PAPOLG is associated with increased transcript expression levels in transformed follicular lymphoma cases. mRNA expression levels of tFL cases with BCL11A (A), PAPOLG (B), PUS10 (C), and USP34 (D) amplifications were compared to those with no amplification of the corresponding gene using probe set values (NCBI GEO dataset: GSE81183) individually and displayed using box-and-whisker plots (t-test, *: P < 0.01). n.s: nonsignificant.

Figure 4

Transcript expression levels of PAPOLG or PUS10 increase in transformed follicular lymphoma tumors compared with those of patient-matched pretransformation tumors. The change in BCL11A (A), PAPOLG (B), PUS10 (C), and USP34 (D) mRNA expression in 12 tFL cases compared to that of pretransformation levels was shown using cDNA microarray clones available in the Lymphochip platform. Clone IDs for each gene are shown near each plot (NCBI GEO accession number: GSE3458).

Figure 5

Transcript expression levels of PAPOLG are significantly higher in follicular lymphoma cases compared to the levels in normal B cells. Expression levels of BCL11A (A), PAPOLG (B), PUS10 (C), and USP34 (D) genes were evaluated in FL cases (n = 63) and normal B cell subsets (n = 6) using the GEP data available (GEO accession number: GSE55267). The expression levels of each gene are shown with box-and-whisker plots (t-test, *: P < 0.05, **: P < 0.01).

Figure 6

The relationship between expression of amplified genes in low- and high-grade follicular lymphoma cases. Transcript expression levels of BCL11A (A), PAPOLG (B), PUS10 (C), and USP34 (D) in low grade (stage 1–2) and high grade (stage 3–3a) FL cases are shown with box-and-whisker plots.