TEMPRANILLO is a direct repressor of the microRNA miR172.

In the age-dependent pathway, microRNA 156 (miR156) is essential for the correct timing of developmental transitions. miR156 negatively regulates several SPL genes, which promote the juvenile-to-adult and floral transitions in part through upregulation of miR172. The transcriptional repressors TEMPRANILLO1 (TEM1) and TEM2 delay flowering in Arabidopsis thaliana at least through direct repression of FLOWERING LOCUS T (FT) and gibberellin biosynthetic genes, and have also been reported to participate in the length of the juvenile phase. Levels of TEM mRNA and miR156 decrease gradually, allowing progression through developmental phases. Given these similarities, we hypothesized that TEMs and the miR156/SPL/miR172 module could act through a common genetic pathway. We analyzed the effect of TEMs on levels of miR156, SPL and miR172, tested binding of TEMs to these genes using chromatin immunoprecipitation and analyzed the genetic interaction between TEMs and miR172. We found that TEMs played a stronger role in the floral transition than in the juvenile-to-adult transition. TEM1 repressed MIR172A, MIR172B and MIR172C expressions and bound in vivo to at least MIR172C sequences. Genetic analyses indicated that TEMs affect the regulation of developmental timing through miR172.

Under a New Light: Regulation of Light-Dependent Pathways by Non-coding RNAs.

The biological relevance of non-protein coding RNAs in the regulation of critical plant processes has been firmly established in recent years. This has been mostly achieved with the discovery and functional characterization of small non-coding RNAs, such as small interfering RNAs and microRNAs (miRNAs). However, recent next-generation sequencing techniques have widened our view of the non-coding RNA world, which now includes long non-coding RNAs (lncRNAs). Small and lncRNAs seem to diverge in their biogenesis and mode of action, but growing evidence highlights their relevance in developmental processes and in responses to particular environmental conditions. Light can affect MIRNA gene transcription, miRNA biogenesis, and RNA-induced silencing complex (RISC) activity, thus controlling not only miRNA accumulation but also their biological function. In addition, miRNAs can mediate several light-regulated processes. In the lncRNA world, few reports are available, but they already indicate a role in the regulation of photomorphogenesis, cotyledon greening, and photoperiod-regulated flowering. In this review, we will discuss how light controls MIRNA gene expression and the accumulation of their mature forms, with a particular emphasis on those miRNAs that respond to different light qualities and are conserved among species. We will also address the role of small non-coding RNAs, particularly miRNAs, and lncRNAs in the regulation of light-dependent pathways. We will mainly focus on the recent progress done in understanding the interconnection between these non-coding RNAs and photomorphogenesis, circadian clock function, and photoperiod-dependent flowering.

TEMPRANILLO Reveals the Mesophyll as Crucial for Epidermal Trichome Formation.

Plant trichomes are defensive specialized epidermal cells. In all accepted models, the epidermis is the layer involved in trichome formation, a process controlled by gibberellins (GAs) in Arabidopsis rosette leaves. Indeed, GA activates a genetic cascade in the epidermis for trichome initiation. Here we report that TEMPRANILLO (TEM) genes negatively control trichome initiation not only from the epidermis but also from the leaf layer underneath the epidermis, the mesophyll. Plants over-expressing or reducing TEM specifically in the mesophyll, display lower or higher trichome numbers, respectively. We surprisingly found that fluorescently labeled GA3 accumulates exclusively in the mesophyll of leaves, but not in the epidermis, and that TEM reduces its accumulation and the expression of several newly identified GA transporters. This strongly suggests that TEM plays an essential role, not only in GA biosynthesis, but also in regulating GA distribution in the mesophyll, which in turn directs epidermal trichome formation. Moreover, we show that TEM also acts as a link between GA and cytokinin signaling in the epidermis by negatively regulating downstream genes of both trichome formation pathways. Overall, these results call for a re-evaluation of the present theories of trichome formation as they reveal mesophyll essential during epidermal trichome initiation.

SHORT VEGETATIVE PHASE Up-Regulates TEMPRANILLO2 Floral Repressor at Low Ambient Temperatures.

Plants integrate day length and ambient temperature to determine the optimal timing for developmental transitions. In Arabidopsis (Arabidopsis thaliana), the floral integrator FLOWERING LOCUS T (FT) and its closest homolog TWIN SISTER OF FT promote flowering in response to their activator CONSTANS under long-day inductive conditions. Low ambient temperature (16°C) delays flowering, even under inductive photoperiods, through repression of FT, revealing the importance of floral repressors acting at low temperatures. Previously, we have reported that the floral repressors TEMPRANILLO (TEM; TEM1 and TEM2) control flowering time through direct regulation of FT at 22°C. Here, we show that tem mutants are less sensitive than the wild type to changes in ambient growth temperature, indicating that TEM genes may play a role in floral repression at 16°C. Moreover, we have found that TEM2 directly represses the expression of FT and TWIN SISTER OF FT at 16°C. In addition, the floral repressor SHORT VEGETATIVE PHASE (SVP) directly regulates TEM2 but not TEM1 expression at 16°C. Flowering time analyses of svp tem mutants indicate that TEM may act in the same genetic pathway as SVP to repress flowering at 22°C but that SVP and TEM are partially independent at 16°C. Thus, TEM2 partially mediates the temperature-dependent function of SVP at low temperatures. Taken together, our results indicate that TEM genes are also able to repress flowering at low ambient temperatures under inductive long-day conditions.

RAV genes: regulation of floral induction and beyond.

BACKGROUND: Transcription factors of the RAV (RELATED TO ABI3 AND VP1) family are plant-specific and possess two DNA-binding domains. In Arabidopsis thaliana, the family comprises six members, including TEMPRANILLO 1 (TEM1) and TEM2. Arabidopsis RAV1 and TEM1 have been shown to bind bipartite DNA sequences, with the consensus motif C(A/C/G)ACA(N)2-8(C/A/T)ACCTG. Through direct binding to DNA, RAV proteins act as transcriptional repressors, probably in complexes with other co-repressors. SCOPE AND CONCLUSIONS: In this review, a summary is given of current knowledge of the regulation and function of RAV genes in diverse plant species, paying particular attention to their roles in the control of flowering in arabidopsis. TEM1 and TEM2 delay flowering by repressing the production of two florigenic molecules, FLOWERING LOCUS T (FT) and gibberellins. In this way, TEM1 and TEM2 prevent precocious flowering and postpone floral induction until the plant has accumulated enough reserves or has reached a growth stage that ensures survival of the progeny. Recent results indicate that TEM1 and TEM2 are regulated by genes acting in several flowering pathways, suggesting that TEMs may integrate information from diverse pathways. However, flowering is not the only process controlled by RAV proteins. Family members are involved in other aspects of plant development, such as bud outgrowth in trees and leaf senescence, and possibly in general growth regulation. In addition, they respond to pathogen infections and abiotic stresses, including cold, dehydration, high salinity and osmotic stress.

A critical appraisal of phloem-mobile signals involved in tuber induction.

The identification of FLOWERING LOCUS T (FT) and several FT homologs as phloem-mobile proteins that regulate flowering has sparked the search for additional homologs involved in the long-distance regulation of other developmental processes. Given that flowering and tuber induction share regulatory pathways, the quest for long-distance tuberization signals has been further stimulated. Several tuberization regulators have been proposed as mobile molecules, including the FT family protein StSP6A, the plant growth regulators gibberellins and the microRNA miR172. Although some of these hypotheses are attractive and plausible, evidence that these molecules are transmissible in potato has yet to be obtained. Two mRNAs encoding transcription factors, StBEL5 and POTATO HOMEOBOX 1 (POTH1), are mobile and correlate with tuber induction. However, evidence that StBEL5 or POTH1 are required for tuberization is not available yet. Therefore, there are several good candidates for long-distance molecules in the tuberization process. Further research should test their role as systemic tuberization signals.

"And yet it moves": cell-to-cell and long-distance signaling by plant microRNAs.

MicroRNAs (miRNAs) are key regulators of numerous genes in many eukaryotes. Some plant miRNAs are involved in developmental and physiological processes that require intercellular or inter-organ signaling. Movement of other small RNAs within plants has been established. Recent findings also demonstrate intercellular signaling by miRNAs and strongly support that a subset of these regulatory molecules move from one cell to another or over long distances. Phloem exudates contain diverse miRNAs and at least two of them, involved in responses to nutrient availability, are transmitted through grafts, indicating long-distance movement. Two miRNAs that regulate developmental processes are present in cells outside their domains of expression. Several results strongly support that one of them moves from cell to cell. Research on a mutant affected in plasmodesmata trafficking indicates that these intercellular channels are required for transmission of miRNA activity to adjacent cells. Moreover, ARGONAUTE proteins might be involved in the regulation of miRNA trafficking. Hypothesis on the features and mechanisms that may determine miRNA mobility are presented. Future challenges include identifying other mobile miRNAs; demonstrating that miRNA movement is required for non-cell autonomous action; and characterizing the mechanisms of translocation and genetic pathways that regulate miRNA movement.

Potato CONSTANS is involved in photoperiodic tuberization in a graft-transmissible manner.

CONSTANS (CO) is involved in the photoperiodic control of plant developmental processes, including flowering in several species and seasonal growth cessation and bud set in trees. It has been proposed that CO could also affect the day-length regulation of tuber induction in Solanum tuberosum (potato), a plant of great agricultural relevance. To address this question, we examined the role of CO in potato. A potato CO-like gene, StCO, was identified and found to be highly similar to a previously reported potato gene of unknown function. Potato plants overexpressing StCO tuberized later than wild-type plants under a weakly inductive photoperiod. StCO silencing promoted tuberization under both repressive and weakly inductive photoperiods, but did not have any effect under strongly inductive short days, demonstrating that StCO represses tuberization in a photoperiod-dependent manner. The effect of StCO on tuber induction was transmitted through grafts. In addition, StCO affected the mRNA levels of StBEL5 - a tuberization promoter, the mRNA of which moves long distances in potato plants - and StFT/StSP6A, a protein highly similar to FLOWERING LOCUS T (FT), which is a key component of systemic flowering signals in other species. We also found that StFT/StSP6A transcript levels correlate with the induction of tuber formation in wild-type plants. These results show that StCO plays an important role in photoperiodic tuberization and, together with the recent demonstration that StFT/StSP6A promotes tuberization, indicate that the CO/FT module participates in controlling this process. Moreover, they support the notion that StCO is involved in the expression of long-distance regulatory signals in potato, as CO does in other species.

Graft-transmissible induction of potato tuberization by the microRNA miR172.

The photoreceptor phytochrome B (PHYB) and the homeodomain protein BEL5 are involved in the response of potato tuber induction to the photoperiod. However, whether they act in the same tuberization pathway is unknown. Here we show the effect of a microRNA, miR172, on this developmental event. miR172 levels are higher under tuber-inducing short days than under non-inductive long days and are upregulated in stolons at the onset of tuberization. Overexpression of this microRNA in potato promotes flowering, accelerates tuberization under moderately inductive photoperiods and triggers tuber formation under long days. In plants with a reduced abundance of PHYB, which tuberize under long days, both BEL5 mRNA and miR172 levels are reduced in leaves and increased in stolons. This, together with the presence of miR172 in vascular bundles and the graft transmissibility of its effect on tuberization, indicates that either miR172 might be mobile or it regulates long-distance signals to induce tuberization. Consistent with this, plants overexpressing miR172 show increased levels of BEL5 mRNA, which has been reported to be transmissible through grafts. Furthermore, we identify an APETALA2-like mRNA containing a miR172 binding site, which is downregulated in plants overexpressing miR172 and plants in which PHYB is silenced. Altogether, our results suggest that miR172 probably acts downstream of the tuberization repressor PHYB and upstream of the tuberization promoter BEL5 and allow us to propose a model for the control of tuberization by PHYB, miR172 and BEL5.

Long-range signalling in plant reproductive development.

Animals and plants produce regulatory signals at specific places of their bodies, in order to regulate developmental events which take place at a distance. Plants use this mechanism to adjust their development to the changing environment. Flowering and tuber formation are controlled by signals generated in the leaves that travel throughout the plant to reach their target tissues: the shoot apical meristem for flowering and the underground stolons for tuberization. Although the existence of these long-distance plant messengers was postulated almost seventy years ago, their chemical nature is still not clear. These leaf-derived signals are graft-transmissible and move through the plant vascular system. Presumably they are very similar or even identical for flowering and tuberization and common to most plant species. It is generally accepted that their composition is complex and includes positive and negative regulators. Many different substances, including classical plant hormones and metabolites have been postulated to be components of these mobile signals, but conclusive evidence of this is still lacking. Recent work has positioned these signals within the genetic network that regulates flowering time and suggests roles for specific genes in the generation, transport or response to the signalling molecules. Current knowledge of long-range signalling mechanisms in other physiological and developmental events, together with the finding of common regulators involved in flowering, tuberization and other processes like pathogen and wound responses, should help to establish the biochemical composition of these elusive messenger signals.

CONSTANS acts in the phloem to regulate a systemic signal that induces photoperiodic flowering of Arabidopsis.

Flower development at the shoot apex is initiated in response to environmental cues. Day length is one of the most important of these and is perceived in the leaves. A systemic signal, called the floral stimulus or florigen, is then transmitted from the leaves through the phloem and induces floral development at the shoot apex. Genetic analysis in Arabidopsis identified a pathway of genes required for the initiation of flowering in response to day length. The nuclear zinc-finger protein CONSTANS (CO) plays a central role in this pathway, and in response to long days activates the transcription of FT, which encodes a RAF-kinase-inhibitor-like protein. We show using grafting approaches that CO acts non-cell autonomously to trigger flowering. Although CO is expressed widely, its misexpression from phloem-specific promoters, but not from meristem-specific promoters, is sufficient to induce early flowering and complement the co mutation. The mechanism by which CO triggers flowering from the phloem involves the cell-autonomous activation of FT expression. Genetic approaches indicate that CO activates flowering through both FT-dependent and FT-independent processes, whereas FT acts both in the phloem and the meristem to trigger flowering. We propose that, partly through the activation of FT, CO regulates the synthesis or transport of a systemic flowering signal, thereby positioning this signal within the established hierarchy of regulatory proteins that controls flowering.