Abstract
ABSTRACThox genes are found as clusters in the genome in most bilaterians. The order of genes in the cluster is supposed to be correlated with the site of expression along the anterior-posterior body axis and the timing of expression during development, and these correlations are called spatial and temporal collinearity, respectively. Here we studied the expression dynamics of all hox genes of the diploid species Xenopus tropicalis in four Hox clusters (A–D) by analyzing high-temporal-resolution RNA-seq databases and the results showed that temporal collinearity is not supported, which is consistent with our previous data from allotetraploid Xenopus laevis. Because the temporal collinearity hypothesis implicitly assumes the collinear order of gene activation, not mRNA accumulation, we determined for the first time the timing of when new transcripts of hox genes are produced, by detecting pre-spliced RNA in whole embryos with reverse transcription and quantitative PCR (RT-qPCR) for all hoxa genes as well as several selected hoxb, hoxc and hoxd genes. Our analyses showed that, coinciding with the RNA-seq results, hoxa genes started to be transcribed in a non-sequential order, and found that multiple genes start expression almost simultaneously or more posterior genes could be expressed earlier than anterior ones. This tendency was also found in hoxb and hoxc genes. These results suggest that temporal collinearity of hox genes is not held during early development of Xenopus.
Highlights
IntroductionWe characterized developmental expression patterns of the whole set of X. laevis hox genes using RNA-seq which measures the amount of mature transcripts, and found that, while a subset of PG1 genes (hoxa.L, hoxb.S and hoxd.L/S) are expressed early, there was no temporal collinearity in genes belonging to PGs 2 through 10 in any of the individual clusters, though the result with hoxd genes was not conclusive because of very low expression levels in many of them
Accumulation of hox gene transcripts during early development Based on the high-resolution expression profiles obtained by RNA-seq of X. tropicalis developmental stages (Owens et al, 2016), which calculated absolute numbers of transcripts in an embryo, we tried to deduce the order that hox genes are expressed by determining the timing that transcripts reach a certain number
reverse transcription and quantitative PCR (RT-qPCR) method evaluation Because RNA-seq analyses are measurements of how much mRNA is present at a given time or in a tissue, we examined the temporal order of gene activation in a Hox cluster, which can be estimated by detecting newly synthesized transcripts
Summary
We characterized developmental expression patterns of the whole set of X. laevis hox genes using RNA-seq which measures the amount of mature transcripts, and found that, while a subset of PG1 genes (hoxa.L, hoxb.S and hoxd.L/S) are expressed early, there was no temporal collinearity in genes belonging to PGs 2 through 10 in any of the individual clusters, though the result with hoxd genes was not conclusive because of very low expression levels in many of them. This apparent lack of temporal collinearity of accumulation of mature transcripts in X. laevis hox genes might be a result of sub- or neofunctionalization causing differences between homeologs, or because temporal collinearity of de novo transcription is just hindered by looking at the accumulation of mature transcripts
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