ADA2b and GCN5 Affect Cytokinin Signaling by Modulating Histone Acetylation and Gene Expression during Root Growth of Arabidopsis thaliana.

In Arabidopsis thaliana, the histone acetyltransferase GCN5 and the associated coactivator ADA2b regulate root growth and affect gene expression. The cytokinin signaling reporter TCS::GFP was introduced into gcn5-1, ada2b-1, and ada2a-2, as well as the ada2a-2ada2b-1 mutants. The early root growth (4 to 7 days post-germination) was analyzed using cellular and molecular approaches. TCS signal accumulated from the fourth to seventh days of root growth in the wild-type columella cells. In contrast, ada2b-1 and gcn5-1 and ada2a-2ada2b-1 double mutants displayed reduced TCS expression relative to wild type. Gene expression analysis showed that genes associated with cytokinin homeostasis were downregulated in the roots of gcn5-1 and ada2b-1 mutants compared to wild-type plants. H3K14 acetylation was affected in the promoters of cytokinin synthesis and catabolism genes during root growth of Arabidopsis. Therefore, GCN5 and ADA2b are positive regulators of cytokinin signaling during root growth by modulating histone acetylation and the expression of genes involved in cytokinin synthesis and catabolism. Auxin application in the roots of wild-type seedlings increased TCS::GFP expression. In contrast, ada2b and ada2ada2b mutant plants do not show the auxin-induced TCS signal, suggesting that GCN5 and ADA2b are required for the auxin-induced cytokinin signaling in early root growth.

Histone Acetyltransferase GCN5 Affects Auxin Transport during Root Growth by Modulating Histone Acetylation and Gene Expression of PINs.

General Control Non-Derepressible 5 (GCN5) is a histone acetyltransferase that targets multiple genes and is essential for the acetylation of Lysine residues in the N-terminal tail of histone H3 in Arabidopsis. GCN5 interacts with the transcriptional coactivator Alteration/Deficiency in Activation 2b (ADA2b), which enhances its activity functioning in multiprotein complexes, such as the Spt-Ada-Gcn5-Acetyltransferase complex (SAGA). Mutations in GCN5 and ADA2b result in pleiotropic phenotypes, including alterations in the growth of roots. Auxin is known to regulate root development by modulating gene expression patterns. Auxin moves polarly during plant growth via the Pin-formed (PIN) auxin efflux transport proteins. The effect of GCN5 and ADA2b on auxin distribution at different stages of early root growth (4 to 7 days post-germination) was studied using the reporter lines DR5rev::GFP and PIN1::PIN1-GFP. In wild-type plants, auxin efflux transporter PIN1 expression increases from the fourth to the seventh day of root growth. The PIN1 expression was reduced in the roots of gcn5-1 and ada2b-1 compared to the wild type. The expression of PIN1 in ada2b-1 mutants is confined only to the meristematic zone, specifically in the stele cells, whereas it is almost abolished in the elongation zone. Gene expression analysis showed that genes associated with auxin transport, PIN1, PIN3 and PIN4, are downregulated in gcn5-1 and ada2b-1 mutants relative to the wild type. As a result, auxin accumulation was also reduced in gcn5-1 and ada2b-1 compared to wild-type roots. Furthermore, acetylation of Lysine 14 of histone H3 (H3K14) was also affected in the promoter and coding region of PIN1, PIN3 and PIN4 genes during root growth of Arabidopsis in gcn5 mutants. In conclusion, GCN5 acts as a positive regulator of auxin distribution in early root growth by modulating histone H3 acetylation and the expression of auxin efflux transport genes.

A Functional Kinase Is Necessary for Cyclin-Dependent Kinase G1 (CDKG1) to Maintain Fertility at High Ambient Temperature in Arabidopsis.

Maintaining fertility in a fluctuating environment is key to the reproductive success of flowering plants. Meiosis and pollen formation are particularly sensitive to changes in growing conditions, especially temperature. We have previously identified cyclin-dependent kinase G1 (CDKG1) as a master regulator of temperature-dependent meiosis and this may involve the regulation of alternative splicing (AS), including of its own transcript. CDKG1 mRNA can undergo several AS events, potentially producing two protein variants: CDKG1L and CDKG1S, differing in their N-terminal domain which may be involved in co-factor interaction. In leaves, both isoforms have distinct temperature-dependent functions on target mRNA processing, but their role in pollen development is unknown. In the present study, we characterize the role of CDKG1L and CDKG1S in maintaining Arabidopsis fertility. We show that the long (L) form is necessary and sufficient to rescue the fertility defects of the cdkg1-1 mutant, while the short (S) form is unable to rescue fertility. On the other hand, an extra copy of CDKG1L reduces fertility. In addition, mutation of the ATP binding pocket of the kinase indicates that kinase activity is necessary for the function of CDKG1. Kinase mutants of CDKG1L and CDKG1S correctly localize to the cell nucleus and nucleus and cytoplasm, respectively, but are unable to rescue either the fertility or the splicing defects of the cdkg1-1 mutant. Furthermore, we show that there is partial functional overlap between CDKG1 and its paralog CDKG2 that could in part be explained by overlapping gene expression.