Abstract
BackgroundThe 29-member Arabidopsis AHL gene family is classified into three main classes based on nucleotide and protein sequence evolutionary differences. These differences include the presence or absence of introns, type and/or number of conserved AT-hook and PPC domains. AHL gene family members are divided into two phylogenetic clades, Clade-A and Clade-B. A majority of the 29 members remain functionally uncharacterized. Furthermore, the biological significance of the DNA and peptide sequence diversity, observed in the conserved motifs and domains found in the different AHL types, is a subject area that remains largely unexplored.ResultsTransgenic plants overexpressing AtAHL20 flowered later than the wild type under both short and long days. Transcript accumulation analyses showed that 35S:AtAHL20 plants contained reduced FT, TSF, AGL8 and SPL3 mRNA levels. Similarly, overexpression of AtAHL20’s orthologue in Camelina sativa, Arabidopsis’ closely related Brassicaceae family member species, conferred a late-flowering phenotype via suppression of CsFT expression. However, overexpression of an aberrant AtAHL20 gene harboring a missense mutation in the AT-hook domain’s highly conserved R-G-R core motif abolished the late-flowering phenotype. Data from targeted yeast-two-hybrid assays showed that AtAHL20 interacted with itself and several other Clade-A Type-I AHLs which have been previously implicated in flowering-time regulation: AtAHL19, AtAHL22 and AtAHL29.ConclusionWe showed via gain-of-function analysis that AtAHL20 is a negative regulator of FT expression, as well as other downstream flowering time regulating genes. A similar outcome in Camelina sativa transgenic plants overexpressing CsAHL20 suggest that this is a conserved function. Our results demonstrate that AtAHL20 acts as a photoperiod-independent negative regulator of transition to flowering.
Highlights
The 29-member Arabidopsis Adenine Thymine (AT)-HOOK MOTIF CONTAINING NUCLEAR LOCALIZED (AHL) gene family is classified into three main classes based on nucleotide and protein sequence evolutionary differences
Strong global GUS signal was observed in all tissues, including root hairs, suggesting that AtAHL20 is constitutively expressed in seedlings (Fig. 1a). 12-day old pAtAHL20-GUS transgenic plants displayed GUS activity in leaf minor veins and trichomes (Fig. 1b)
Multiple independent transgenic plants overexpressing AtAHL20 displayed a dwarf phenotype (Fig. 2a, b) and a late-flowering phenotype compared to the wild type and ahl20-1/ahl20-2 TDNA insertion lines under both LDs (Fig. 2c, d) and SDs (Fig. 2e, f)
Summary
The 29-member Arabidopsis AHL gene family is classified into three main classes based on nucleotide and protein sequence evolutionary differences These differences include the presence or absence of introns, type and/or number of conserved AT-hook and PPC domains. The 29-member Arabidopsis AT-HOOK MOTIF CONTAINING NUCLEAR LOCALIZED (AHL) gene family is found in all sequenced plant species, ranging from the. Zhao et al [49] showed that sob, a dominant negative mutant carrying a missense allele in the R-G-R core of the AT-hook motif, displayed more dramatic hypocotyl phenotypes compared to the sob esc-8 ahl ahl quadruple mutant Based on these data, a molecular model was proposed where AHLs interact with each other and themselves, as well as other nuclear proteins, such as transcription factors (TFs), to form a “DNA-AHL-TF complex” [49]. It is hypothesized that most AHLs function as complexes, and that mutations in the DNA-binding AThook motif may render that entire complex nonfunctional [49]
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