Exposure to fluorescent light triggers down regulation of genes involved with mitotic progression in Xiphophorus skin

Abstract

We report RNA-Seq results from skin of X. maculatus Jp 163 B after exposure to various doses of "cool white" fluorescent light (FL). We show that FL exposure incites a genetic transcriptional response in skin nearly as great as observed for UVB exposure; however, the gene sets modulated due to exposure to the two light sources are quite different. Known light responsive genes involved in maintaining circadian cycling (e.g., clock, cry2a, cry1b, per1b, per2, per3, and arntl1a) exhibited expected shifts in transcriptional expression upon FL exposure. Exposure to FL also resulted in down-regulated transcription of many genes involved with cell cycle progression (e.g., cdc20, cdc45, cdca7b, plk1, cdk1, ccnb-3, and cdca7a) and chromosome segregation (e.g., cenpe, cenpf, cenpi, cenpk, cenpo, cenpp, and cenpu; cep70; knstrm, kntc, mcm2, mcm5; smc2). In addition, several DNA replication and recombination repair genes (e.g., pola1, pole, rec52, rad54l, rpa1, and parpbp) exhibit reduced expression in FL exposed X. maculatus skin. Some genes down modulated by FL are known to be associated with DNA repair and human diseases (e.g., atm2, brip1, fanc1, fancl, and xrcc4). The overall suppression of genes involved with mitotic progression in the skin of adult fish is consistent with entry into the light phase of the circadian cycle. Current efforts are aimed at determining specific wavelengths that may lead to differential expression among the many genes affected by fluorescent light exposure.

Keywords: Cell cycle progression; Circadian; Cool white light; DNA damage response; Differential gene expression; Fluorescent light; RNA-Seq.

Figure 1

The total number of up- and down-modulated genes in X. maculatus Jp 163 B skin due to UVB (8 kJ) or FL light exposures (20, 40, or 60 min). Within each bar are numbers of genes with increased transcription levels (top bar), or genes exhibiting depressed transcription levels (bottom bar) compared to the skin of unexposed sibling animals. The total number of genes showing modulated transcription (DE genes) are 322 for UVB, 271 for 20 min FL, 282 for 40 min FL, and 274 at 60 min FL.

Figure 2

Venn diagram comparison of DE genes from each FL exposure (20, 40, and 60 min). There are 134 genes in common between the 20 min (49%) and 40 min (48%) time points, 147 genes in common between the 20 min (54%) and 60 min (54%) time points, and 117 genes in common between the 40 min (41%) and 60 min (43%) time points. Among all three exposures, 91 DE genes are shared and expressed similarly (see Table S4).

Figure 3

Relative transcription levels of a set of genes identified as similarly affected by UV or FL exposure. This set of 32 genes include known light responsive genes (which are upregulated) and genes involved in mitosis and intercellular signaling (which are downregulated). For instance, light responsive (ybx2, sox3) and circadian cycle genes (clock, per2) are up regulated while many of down regulated genes play a role in mitosis (aspm, cenp-e, -f, fam64a, kif-14 and -20a, knstrn, kpna2, tacc3, etc.), cell cycling (ccnb1, cdc20), or intercellular signaling (raver1, arhgap19, depdc1a).

Figure 4

Genetic associations of chromosome structure and mitosis genes differentially modulated in at least 2 time points (20, 40, and 60 min FL exposure) in X. maculatus JP 163 B skin. Biological associations are identified through genetic interaction (green lines) and pathways (blue lines). Grey circles are genes predicted through the GeneMania algorithm to be associated with query genes (black circles).

Figure 5

NanoString analysis confirmed the RNA-Seq data for 18 genes in both direction and magnitude (R² = 0.86) following 20 (red), 40 (green) and 60 (blue) min FL exposure. Fold changes are represented as the average change of 2 biological replicates determined independently for each target plotted and at each exposure.

Figure 6

RNA-Seq validation (20 min FL- light grey; 40 min FL- dark grey) was performed on seven targets using qRT-PCR following 20 (red) and 40 (green) min FL exposure. Mean fold change relative to an unexposed sample was calculated using the ΔΔCT method with an 18S rRNA internal standard for 3 technical replicates.