Effects of TSA, NaB, Aza in Lactuca sativa L. protoplasts and effect of TSA in Nicotiana benthamiana protoplasts on cell division and callus formation

Abstract

Whole-plant regeneration via plant tissue culture is a complex process regulated by several genetic and environmental conditions in plant cell cultures. Recently, epigenetic regulation has been reported to play an important role in plant cell differentiation and establishment of pluripotency. Herein, we tested the effects of chemicals, which interfere with epigenetic regulation, on the plant regeneration from mesophyll protoplasts of lettuce. The used chemicals were histone deacetylase inhibitors trichostatin A (TSA) and sodium butyrate (NaB), and the DNA methyltransferase inhibitor azacytidine (Aza). All three chemicals increased cell division, micro-callus formation and callus proliferation in lettuce protoplasts. Cell division increased by more than 20% with an optimal treatment of the three chemicals. In addition, substantial increase in the callus proliferation rates was observed. In addition, TSA enhances cell division and adventitious shoot formation in the protoplast culture of Nicotiana benthamiana. The regenerated tobacco plants from TSA-treated protoplasts did not show morphological changes similar to the control. TSA increased histone H3 acetylation levels and affected the expression of CDK, CYCD3-1, and WUS in tobacco protoplasts. Thus, we investigated the effect of TSA, NaB, and Aza on Lactuca sativa L. protoplasts and the effect of TSA on cell division and callus formation in Nicotiana benthamiana protoplasts, which facilitates plant regeneration from mesophyll protoplasts. Furthermore, these chemicals can be directly applied as media additives for efficient plant regeneration and crop improvement in various plant species.

Multiscale chromatin dynamics and high entropy in plant iPSC ancestors

“…Next, we tested the influence of trichostatin A (TSA), a compound known to inhibit histone deacetylases and increase histone acetylation ( Yoshida et al, 1995 ). TSA treatment was shown to enhance the regenerative competence of lettuce and Nicotiana protoplast cultures, notably inducing a higher rate of division starting from day 5 ( Choi et al, 2023 ). We rationalised that this drug could possibly accelerate chromatin decondensation or alter the chromatin feature dynamics that were detected with our approach.…”

Unraveling Epigenetic Changes in A. thaliana Calli: Impact of HDAC Inhibitors

“…Data on the effect of NaB on callus formation are scarce. Recently, the positive effects of NaB, TSA, and azacytidine, an inhibitor of DNA methyltransferases, on lettuce callus formation and propagation were presented [ 39 ]. Although in our experiments NaB’s influence on callus formation was less pronounced compared with TSA, NaB clearly sped up the formation of microcalli on the shoot part of the seedlings ( Figure 1 A).…”

The improvement of the in vitro plant regeneration in barley with the epigenetic modifier of histone acetylation, trichostatin A

“…The genotype-dependent results of TSA treatment were also observed in B. rapa (Zhang et al 2016 ) and wheat (Jiang et al 2017 ; Bie et al 2020 ). Besides genotype, the effect of TSA on plant regeneration depends on treatment conditions such as concentration and treatment time (Castillo et al 2020 ; Martínez et al 2021 ; Choi et al 2023 ). We tested the effect of TSA at a wide range of concentrations (1.0–7.5 μM) and durations of treatment (1, 2, and 4 weeks) and indicated the dose of 7.5 μM of TSA for 4 weeks to be effective in stimulation of plant regeneration in some barley genotypes.…”