The histone replacement gene His4r is involved in heat stress induced chromatin rearrangement

His4r is the only known variant of histone H4 in Drosophila. It is encoded by the His4r single-copy gene that is located outside of the histone gene cluster and expressed in a different pattern than H4, although the encoded polypeptides are identical. We generated a null mutant (His4rΔ42) which is homozygous viable and fertile without any apparent morphological defects. Heterozygous His4rΔ42 is a mild suppressor of position-effect variegation, suggesting that His4r has a role in the formation or maintenance of condensed chromatin. Under standard conditions loss of His4r has a modest effect on gene expression. Upon heat-stress the induction of the Heat shock protein (HSP) genes Hsp27and Hsp68 is stronger in His4rΔ42 mutants with concordantly increased survival rate. Analysis of chromatin accessibility after heat shock at aHsp27 regulatory region showed less condensed chromatin in the absence of His4r while there was no difference at the gene body.

Figure 1

Fig: Generation and validation of the His4rΔ42 deletion mutant. (A) The genomic region of His4r on chromosome arm 3R with transcript models, primers and the extent of the deletion is shown. On the His4r transcript models coding sequences are represented by black bars, untranslated exonic regions by empty bars, and introns by solid lines. Adjacent ends of neighboring genes (put and CG7886) are shown with grey bars. Empty arrowhead indicates the position of the P{EPgy2}His4rEY06726 transposon in the His4r 5′-UTR, brackets show the breakpoints of the His4rΔ42 deletion. Position of PCR primers are marked by black arrows. (B) PCR reactions on homozygous His4rΔ42 template with His4rFseq and His4r-E3CR primers did not give a PCR product, with His4rFseq and His4rgR primers the length of the product was approximately 700 bp shorter than on wild-type control template. PCR reactions on His4rrev5templates resulted in amplicons similar to wild-type. (C) His4r transcript levels were similar in Oregon-R (wild-type) and in His4rrev5revertant flies (P > 0.05), while in His4rΔ42 mutants His4r mRNA was not present (P = 2.69077 × 10−5, t-test). The bars show the mean of normalized RT-qPCR measurements relative to a template dilution series, error bars represent standard error, n = 3.

 

Interestingly, preconditioning before heat shock led to increased chromatin accessibility, HSP gene transcription and survival rate in control flies while it did not cause notable changes in His4rΔ42. Thus, our results suggest that His4r might play a role in fine tuning chromatin structure at inducible gene promoters upon environmental stress conditions.

 

Faragó, A., Ürmösi, A., Farkas, A. et al. The histone replacement geneHis4r is involved in heat stress induced chromatin rearrangement. Sci Rep 11, 4878 (2021). https://doi.org/10.1038/s41598-021-84413-4

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