Root-knot nematodes induce gall formation by recruiting developmental pathways of post-embryonic organogenesis and regeneration to promote transient pluripotency.

Root-knot nematodes (RKNs; Meloidogyne spp.) induce new post-embryogenic organs within the roots (galls) where they stablish and differentiate nematode feeding cells, giant cells (GCs). The developmental programmes and functional genes involved remain poorly defined. Arabidopsis root apical meristem (RAM), lateral root (LR) and callus marker lines, SHORT-ROOT/SHR, SCARECROW/SCR, SCHIZORIZA/SCZ, WUSCHEL-RELATED-HOMEOBOX-5/WOX5, AUXIN-RESPONSIVE-FACTOR-5/ARF5, ARABIDOPSIS-HISTIDINE PHOSPHOTRANSFER-PROTEIN-6/AHP6, GATA-TRANSCRIPTION FACTOR-23/GATA23 and S-PHASE-KINASE-ASSOCIATED-PROTEIN2B/SKP2B, were analysed for nematode-dependent expression. Their corresponding loss-of-function lines, including those for LR upstream regulators, SOLITARY ROOT/SLR/IAA14, BONDELOS/BDL/IAA12 and INDOLE-3-ACETIC-ACID-INDUCIBLE-28/IAA28, were tested for RKN resistance/tolerance. LR genes, for example ARF5 (key factor for root stem-cell niche regeneration), GATA23 (which specifies pluripotent founder cells) and AHP6 (cytokinin-signalling-inhibitor regulating pericycle cell-divisions orientation), show a crucial function during gall formation. RKNs do not compromise the number of founder cells or LR primordia but locally induce gall formation possibly by tuning the auxin/cytokinin balance in which AHP6 might be necessary. Key RAM marker genes were induced and functional in galls. Therefore, the activation of plant developmental programmes promoting transient-pluripotency/stemness leads to the generation of quiescent-centre and meristematic-like cell identities within the vascular cylinder of galls. Nematodes enlist developmental pathways of new organogenesis and/or root regeneration in the vascular cells of galls. This should determine meristematic cell identities with sufficient transient pluripotency for gall organogenesis.

Regulation of Hormonal Control, Cell Reprogramming, and Patterning during De Novo Root Organogenesis.

Body regeneration through formation of new organs is a major question in developmental biology. We investigated de novo root formation using whole leaves of Arabidopsis (Arabidopsis thaliana). Our results show that local cytokinin biosynthesis and auxin biosynthesis in the leaf blade followed by auxin long-distance transport to the petiole leads to proliferation of J0121-marked xylem-associated tissues and others through signaling of INDOLE-3-ACETIC ACID INDUCIBLE28 (IAA28), CRANE (IAA18), WOODEN LEG, and ARABIDOPSIS RESPONSE REGULATORS1 (ARR1), ARR10, and ARR12. Vasculature proliferation also involves the cell cycle regulator KIP-RELATED PROTEIN2 and ABERRANT LATERAL ROOT FORMATION4, resulting in a mass of cells with rooting competence that resembles callus formation. Endogenous callus formation precedes specification of postembryonic root founder cells, from which roots are initiated through the activity of SHORT-ROOT, PLETHORA1 (PLT1), and PLT2. Primordia initiation is blocked in shr plt1 plt2 mutant. Stem cell regulators SCHIZORIZA, JACKDAW, BLUEJAY, and SCARECROW also participate in root initiation and are required to pattern the new organ, as mutants show disorganized and reduced number of layers and tissue initials resulting in reduced rooting. Our work provides an organ regeneration model through de novo root formation, stating key stages and the primary pathways involved.

Whole genome duplications in plants: an overview from Arabidopsis.

Polyploidy is a common event in plants that involves the acquisition of more than two complete sets of chromosomes. Allopolyploidy originates from interspecies hybrids while autopolyploidy originates from intraspecies whole genome duplication (WGD) events. In spite of inconveniences derived from chromosomic rearrangement during polyploidization, natural plant polyploids species often exhibit improved growth vigour and adaptation to adverse environments, conferring evolutionary advantages. These advantages have also been incorporated into crop breeding programmes. Many tetraploid crops show increased stress tolerance, although the molecular mechanisms underlying these different adaptation abilities are poorly known. Understanding the physiological, cellular, and molecular mechanisms coupled to WGD, in both allo- and autopolyploidy, is a major challenge. Over the last few years, several studies, many of them in Arabidopsis, are shedding light on the basis of genetic, genomic, and epigenomic changes linked to WGD. In this review we summarize and discuss the latest advances made in Arabidopsis polyploidy, but also in other agronomic plant species.

Identification of a protein network interacting with TdRF1, a wheat RING ubiquitin ligase with a protective role against cellular dehydration.

Plants exploit ubiquitination to modulate the proteome with the final aim to ensure environmental adaptation and developmental plasticity. Ubiquitination targets are specifically driven to degradation through the action of E3 ubiquitin ligases. Genetic analyses have indicated wide functions of ubiquitination in plant life; nevertheless, despite the large number of predicted E3s, only a few of them have been characterized so far, and only a few ubiquitination targets are known. In this work, we characterized durum wheat (Triticum durum) RING Finger1 (TdRF1) as a durum wheat nuclear ubiquitin ligase. Moreover, its barley (Hordeum vulgare) homolog was shown to protect cells from dehydration stress. A protein network interacting with TdRF1 has been defined. The transcription factor WHEAT BEL1-TYPE HOMEODOMAIN1 (WBLH1) was degraded in a TdRF1-dependent manner through the 26S proteasome in vivo, the mitogen-activated protein kinase TdWNK5 [for Triticum durum WITH NO LYSINE (K)5] was able to phosphorylate TdRF1 in vitro, and the RING-finger protein WHEAT VIVIPAROUS-INTERACTING PROTEIN2 (WVIP2) was shown to have a strong E3 ligase activity. The genes coding for the TdRF1 interactors were all responsive to cold and/or dehydration stress, and a negative regulative function in dehydration tolerance was observed for the barley homolog of WVIP2. A role in the control of plant development was previously known, or predictable based on homology, for wheat BEL1-type homeodomain1(WBLH1). Thus, TdRF1 E3 ligase might act regulating the response to abiotic stress and remodeling plant development in response to environmental constraints.

The cold-induced factor CBF3 mediates root stem cell activity, regeneration, and developmental responses to cold.

Plant growth and development involve the specification and regeneration of stem cell niches (SCNs). Although plants are exposed to disparate environmental conditions, how environmental cues affect developmental programs and stem cells is not well understood. Root stem cells are accommodated in meristems in SCNs around the quiescent center (QC), which maintains their activity. Using a combination of genetics and confocal microscopy to trace morphological defects and correlate them with changes in gene expression and protein levels, we show that the cold-induced transcription factor (TF) C-REPEAT BINDING FACTOR 3 (CBF3), which has previously been associated with cold acclimation, regulates root development, stem cell activity, and regeneration. CBF3 is integrated into the SHORT-ROOT (SHR) regulatory network, forming a feedback loop that maintains SHR expression. CBF3 is primarily expressed in the root endodermis, whereas the CBF3 protein is localized to other meristematic tissues, including root SCNs. Complementation of cbf3-1 using a wild-type CBF3 gene and a CBF3 fusion with reduced mobility show that CBF3 movement capacity is required for SCN patterning and regulates root growth. Notably, cold induces CBF3, affecting QC activity. Furthermore, exposure to moderate cold around 10°C-12°C promotes root regeneration and QC respecification in a CBF3-dependent manner during the recuperation period. By contrast, CBF3 does not appear to regulate stem cell survival, which has been associated with recuperation from more acute cold (∼4°C). We propose a role for CBF3 in mediating the molecular interrelationships among the cold response, stem cell activity, and development.

An auxin-regulable oscillatory circuit drives the root clock in Arabidopsis.

In Arabidopsis, the root clock regulates the spacing of lateral organs along the primary root through oscillating gene expression. The core molecular mechanism that drives the root clock periodicity and how it is modified by exogenous cues such as auxin and gravity remain unknown. We identified the key elements of the oscillator (AUXIN RESPONSE FACTOR 7, its auxin-sensitive inhibitor IAA18/POTENT, and auxin) that form a negative regulatory loop circuit in the oscillation zone. Through multilevel computer modeling fitted to experimental data, we explain how gene expression oscillations coordinate with cell division and growth to create the periodic pattern of organ spacing. Furthermore, gravistimulation experiments based on the model predictions show that external auxin stimuli can lead to entrainment of the root clock. Our work demonstrates the mechanism underlying a robust biological clock and how it can respond to external stimuli.

Degradation of the cyclin-dependent kinase inhibitor KRP1 is regulated by two different ubiquitin E3 ligases.

In animals and fungi, a group of proteins called the cyclin-dependent kinase inhibitors play a key role in cell cycle regulation. However, comparatively little is known about the role of these proteins in plant cell cycle regulation. To gain insight into the mechanisms by which the plant cell cycle is regulated, we studied the cyclin-dependent kinase inhibitor KRP1 in Arabidopsis. KRP1 interacts with the CDKA;1/CYCD2;1 complex in planta and functions in the G1-S transition of the cell cycle. Furthermore, we show that KRP1 is a likely target of the ubiquitin/proteasome pathway. Two different ubiquitin protein ligases, SCF(SKP2) and the RING protein RKP, contribute to its degradation. These results suggest that SCF(SKP2b) and RPK play an important role in the cell cycle through regulating KRP1 protein turnover.

Polyamines interfere with protein ubiquitylation and cause depletion of intracellular amino acids: a possible mechanism for cell growth inhibition.

Spermidine is a polyamine present in eukaryotes with essential functions in protein synthesis. At high concentrations spermidine and norspermidine inhibit growth by unknown mechanisms. Transcriptomic analysis of the effect of norspermidine on the plant Arabidopsis thaliana indicates upregulation of the response to heat stress and denatured proteins. Accordingly, these polyamines inhibit protein ubiquitylation, both in vivo (in yeast, Arabidopsis, and human Hela cells) and in vitro (with recombinant ubiquitin ligase). This interferes with protein degradation by the proteasome, a situation known to deplete cells of amino acids. Norspermidine treatment of yeast cells induces amino acid depletion, and supplementation of media with amino acids counteracts growth inhibition and cellular amino acid depletion but not inhibition of protein polyubiquitylation.

A Novel Proteomic Analysis of the Modifications Induced by High Hydrostatic Pressure on Hazelnut Water-Soluble Proteins.

Food allergies to hazelnut represent an important health problem in industrialized countries because of their high prevalence and severity. Food allergenicity can be changed by several processing procedures since food proteins may undergo modifications which could alter immunoreactivity. High-hydrostatic pressure (HHP) is an emerging processing technology used to develop novel and high-quality foods. The effect of HHP on allergenicity is currently being investigated through changes in protein structure. Our aim is to evaluate the effect of HHP on the protein profile of hazelnut immunoreactive extracts by comparative proteomic analysis with ProteomeLab PF-2D liquid chromatography and mass spectrometry. This protein fractionation method resolves proteins by isoelectric point and hydrophobicity in the first and second dimension, respectively. Second dimension chromatogram analyses show that some protein peaks present in unpressurized hazelnut must be unsolubilized and are not present in HHP-treated hazelnut extracts. Our results show that HHP treatment at low temperature induced marked changes on hazelnut water-soluble protein profile.

Regulation of the new Arabidopsis imprinted gene AtBMI1C requires the interplay of different epigenetic mechanisms.

Recently, it has been shown that plants contain homologs to the animal Polycomb repressive complex 1 (PRC1) components BMI1 and RING1A/B. In Arabidopsis, there are three BMI1-like genes, two of which, AtBMI1A and B, are required during post-embryonic plant growth to repress embryonic traits and allow cell differentiation. However, little is known about the third BMI1-like gene, AtBMI1C. In this work, we show that AtBMI1C is only expressed during endosperm and stamen development. AtBMI1C is an imprinted gene expressed from the maternal allele in the endosperm but biallelically expressed in stamen. We found that the characteristic expression pattern of AtBMI1C is the result of a complex epigenetic regulation that involves CG DNA methylation, RNA-directed non-CG DNA methylation (RdDM), and PcG activity. Our results show the orchestrated interplay of different epigenetic mechanisms in regulating gene expression throughout development, shedding light on the current hypotheses for the origin and mechanism of imprinting in plant endosperm.

INCURVATA2 encodes the catalytic subunit of DNA Polymerase alpha and interacts with genes involved in chromatin-mediated cellular memory in Arabidopsis thaliana.

Cell type-specific gene expression patterns are maintained by the stable inheritance of transcriptional states through mitosis, requiring the action of multiprotein complexes that remodel chromatin structure. Genetic and molecular interactions between chromatin remodeling factors and components of the DNA replication machinery have been identified in Schizosaccharomyces pombe, indicating that some epigenetic marks are replicated simultaneously to DNA with the participation of the DNA replication complexes. This model of epigenetic inheritance might be extended to the plant kingdom, as we report here with the positional cloning and characterization of INCURVATA2 (ICU2), which encodes the putative catalytic subunit of the DNA polymerase alpha of Arabidopsis thaliana. The strong icu2-2 and icu2-3 insertional alleles caused fully penetrant zygotic lethality when homozygous and incompletely penetrant gametophytic lethality, probably because of loss of DNA polymerase activity. The weak icu2-1 allele carried a point mutation and caused early flowering, leaf incurvature, and homeotic transformations of sepals into carpels and of petals into stamens. Further genetic analyses indicated that ICU2 interacts with TERMINAL FLOWER2, the ortholog of HETEROCHROMATIN PROTEIN1 of animals and yeasts, and with the Polycomb group (PcG) gene CURLY LEAF. Another PcG gene, EMBRYONIC FLOWER2, was found to be epistatic to ICU2. Quantitative RT-PCR analyses indicated that a number of regulatory genes were derepressed in the icu2-1 mutant, including genes associated with flowering time, floral meristem, and floral organ identity.

The HVE/CAND1 gene is required for the early patterning of leaf venation in Arabidopsis.

The hemivenata-1 (hve-1) recessive allele was isolated in a search for natural variations in the leaf venation pattern of Arabidopsis thaliana, where it was seen to cause extremely simple venation in vegetative leaves and cotyledons, increased shoot branching, and reduced root waving and fertility, traits that are reminiscent of some mutants deficient in auxin signaling. Reduced sensitivity to exogenous auxin was found in the hve-1 mutant, which otherwise displayed a wild-type response to auxin transport inhibitors. The HVE gene was positionally cloned and found to encode a CAND1 protein. The hve-1 mutation caused mis-splicing of the transcripts of the HVE/CAND1 gene and a vein phenotype indistinguishable from that of hve-2 and hve-3, two putatively null T-DNA alleles. Inflorescence size and fertility were more affected by hve-2 and hve-3, suggesting that hve-1 is hypomorphic. The simple venation pattern of hve plants seems to arise from an early patterning defect. We found that HVE/CAND1 binds to CULLIN1, and that the venation patterns of axr1 and hve mutants are similar, which suggest that ubiquitin-mediated auxin signaling is required for venation patterning in laminar organs, the only exception being cauline leaves. Our analyses of double mutant and transgenic plants indicated that auxin transport and perception act independently to pattern leaf veins, and that the HVE/CAND1 gene acts upstream of ATHB-8 at least in higher order veins, in a pathway that involves AXR1, but not LOP1, PIN1, CVP1 or CVP2.

Arabidopsis E2Fc functions in cell division and is degraded by the ubiquitin-SCF(AtSKP2) pathway in response to light.

Selective ubiquitin-mediated proteolysis through the cell cycle controls the availability, and therefore the activity, of several cell proliferation proteins. E2F transcription factors play distinct roles in both proliferating and differentiated cells by regulating gene expression. Here, we report that Arabidopsis AtE2Fc is regulated by a balance between gene expression and ubiquitin-proteasome proteolysis. AtE2Fc degradation implicates the function of the E3 ubiquitin-ligase Skp1, Cullin, F-box (SCF(AtSKP2)) complex and seems to be dependent on cyclin-dependent kinase phosphorylation. In addition, we found that AtE2Fc degradation is triggered by light stimulation of dark-grown seedlings. Interestingly, the auxin response mutant axr1-12, in which RUB1 modification of the SCF component CUL1 is impaired, shows increased AtE2Fc protein levels, suggesting a dysfunction in the control of AtE2Fc stability. Likewise, overexpression of a stable form of the AtE2Fc protein negatively affects cell division and increases cell size. These effects are mediated, at least in part, by downregulating the cell cycle gene AtCDC6. The negative role of AtE2Fc in gene expression is further supported by the fact that AtE2Fc interacts with plant retinoblastoma-related protein, suggesting that AtE2Fc might form part of a repressor complex. We propose that AtE2Fc might play a role in cell division and during the transition from skotomorphogenesis to photomorphogenesis.

Comparación entre la zona facial media y el tercio facial inferior en estudiantes de 19 a 25 años de edad de la Facultad de Estomatología de la UPCH / Comparison of middle facial zone and inferior facial third in 19 to 25 years old dental students

El propósito del presente estudio de corte transversal fue comparar las proporciones faciales de 59 estudiantes para evaluar la dimensión vertical. Las medidas se obtuvieron de acuerdo a la técnica de Goodfriend, conocida también como técnica de Willis. Se marcaron cuatro puntos en los rostros de los sujetos y se procedió a medir las proporciones: tercio facial inferior y zona facial media, con la ayuda de una regla pie de rey. La zona facial media se registró solamente en un momento, mientras que el tercio facial inferior se registró en tres momentos cada una de las medidas: en posición postural y en posición de máxima intercuspidación, anotándose el promedio de cada una de las medidas.