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Topicals and chemical peels are the standard of care for management of facial hyperpigmentation. However, traditional therapies have come under recent scrutiny, such as topical hydroquinone (HQ) has some regulatory restrictions, and high concentration trichloroacetic acid (TCA) peel pose a risk in patients with skin of colour. The objective of our research was to identify, investigate and elucidate the mechanism of action of a novel TCA- and HQ-free professional-use chemical peel to manage common types of facial hyperpigmentation. Using computational modelling and in vitro assays on tyrosinase, we identified proprietary multi-acid synergistic technology (MAST). After a single application on human skin explants, MAST peel was found to be more effective than a commercial HQ peel in inhibiting melanin (histochemical imaging and gene expression). All participants completed the case study (N = 9) without any adverse events. After administration of the MAST peel by a dermatologist, the scoring and VISIA photography reported improvements in hyperpigmentation, texture and erythema, which could be linked to underlying pathophysiological changes in skin after peeling, visualized by non-invasive optical biopsy of face. Using reflectance confocal microscopy (VivaScope®) and multiphoton tomography (MPTflexTM), we observed reduction in melanin, increase in metabolic activity of keratinocytes, and no signs of inflammatory cells after peeling. Subsequent swabbing of the cheek skin found no microbiota dysbiosis resulting from the chemical peel. The strong efficacy with minimum downtime and no adverse events could be linked to the synergistic action of the ingredients in the novel HQ- and TCA-free professional peel technology.

Background:Phytocystatins(PhyCys)actasendogenousregulatorsofcysteineproteases(CysProt) involvedin variousphysiologicalprocesses.Besides,PhyCysareinvolvedinplant reactionstoabioticstresses likedroughtor darknessandhavebeenusedaseffectivemoleculesagainstdifferentpestsandpathogens.ThebarleyPhyCysCysProtsystemisconsideredamodelofprotease-inhibitor regulationofproteinturnover.Thirteenbarleycystatins (HvCPI-1toHvCPI-13)havebeenpreviouslyidentifiedandcharacterized.AmongthemHvCPI-2hasbeenshownto havearelevant roleinplant responsestopathogensandpests,aswellas intheplant responsetodrought. Results:Thepresentworkexploresthemultipleroleof thisbarleyPhyCys inresponsetoboth,bioticandabiotic stresses, focusingontheimpactofsilencingthisgene.HvIcy-2silencinglinesbehavedifferentiallyagainst the phytopathogenicfungusMagnaportheoryzaeandalightdeprivationtreatment.TheinducedexpressionofHvIcy-2 bythefungal stresscorrelatedtoahighersusceptibilityofsilencingHvIcy-2plants. Incontrast, areductioninthe expressionofHvIcy-2andinthecathepsin-Land-Blikeactivities inthesilencingHvIcy-2plantswasnot accompaniedbyapparentphenotypicaldifferenceswithcontrolplants inresponsetolightdeprivation. Conclusion:Theseresultshighlight thespecificityofPhyCys intheresponsestodiverseexternalpromptsaswellas thecomplexityof theregulatoryevents leadingtotheresponsetoaparticularstress.Themechanismof regulation of thesestressresponsesseemstobefocusedinmaintainingthebalanceofCysProtandPhyCys levels,whichis crucial for themodulationofphysiologicalprocesses inducedbybioticorabioticstresses., Ministerio de Economia y Competitividad, Depto. de Bioquímica y Biología Molecular, Fac. de Ciencias Biológicas, TRUE, pub

Co-evolutionary processes in plant–pathogen/herbivore systems indicate that protease inhibitors have a particular value in biotic interactions. However, little is known about the defensive role of their targets, the plant proteases. C1A cysteine proteases are the most abundant enzymes responsible for the proteolytic activity during different processes like germination, development and senescence in plants. To identify and characterize C1A cysteine proteases of barley with a potential role in defense, mRNA and protein expression patterns were analyzed in response to biotics stresses. A barley cysteine protease, HvPap-1, previously related to abiotic stresses and grain germination, was particularly induced by flagellin or chitosan elicitation, and biotic stresses such as the phytopathogenic fungus Magnaporthe oryzae or the phytophagous mite Tetranychus urticae. To elucidate the in vivo participation of this enzyme in defense, transformed barley plants overexpressing or silencing HvPap-1 encoding gene were subjected to M. oryzae infection or T. urticae infestation. Whereas overexpressing plants were less susceptible to the fungus than silencing plants, the opposite behavior occurred to the mite. This unexpected result highlights the complexity of the regulatory events leading to the response to a particular biotic stress., Ministerio de Economia y Competitividad, Depto. de Bioquímica y Biología Molecular, Fac. de Ciencias Biológicas, TRUE, pub

C1A plant cysteine proteases are synthesized as pre-pro-enzymes that need to be processed to become active by the pro-peptide claves off from its cognate enzyme. These pro-sequences play multifunctional roles including the capacity to specifically inhibit their own as well as other C1A protease activities from diverse origin. In this study, it is analysed the potential role of C1A pro-regions from barley as regulators of cysteine proteases in target phytophagous arthropods (coleopteran and acari). The in vitro inhibitory action of these pro-sequences, purified as recombinant proteins, is demonstrated. Moreover, transgenic Arabidopsis plants expressing different fragments of HvPap-1 barley gene containing the pro-peptide sequence were generated and the acaricide function was confirmed by bioassays conducted with the two-spotted spider mite Tetranychus urticae. Feeding trials resulted in a significant reduction of leaf damage in the transgenic lines expressing the pro-peptide in comparison to non-transformed control and strongly correlated with an increase in mite mortality. Additionally, the analysis of the expression levels of a selection of potential mite targets (proteases and protease inhibitors) revealed a mite strategy to counteract the inhibitory activity produced by the C1A barley pro-prodomain. These findings demonstrate that pro-peptides can control mite pests and could be applied as defence proteins in biotechnological systems., Ministerio de Economía y Competitividad (MINECO), Depto. de Bioquímica y Biología Molecular, Fac. de Ciencias Biológicas, TRUE, pub

Background: Peptidases are key proteins involved in essential plant physiological processes. Although protein peptidase inhibitors are essential molecules that modulate peptidase activity, their global presence in different plant species remains still unknown. Comparative genomic analyses are powerful tools to get advanced knowledge into the presence and evolution of both, peptidases and their inhibitors across the Viridiplantae kingdom. Results: A genomic comparative analysis of peptidase inhibitors and several groups of peptidases in representative species of different plant taxonomic groups has been performed. The results point out: i) clade-specific presence is common to many families of peptidase inhibitors, being some families present in most land plants; ii) variability is a widespread feature for peptidase inhibitory families, with abundant species-specific (or clade-specific) gene family proliferations; iii) peptidases are more conserved in different plant clades, being C1A papain and S8 subtilisin families present in all species analyzed; and iv) a moderate correlation among peptidases and their inhibitors suggests that inhibitors proliferated to control both endogenous and exogenous peptidases. Conclusions: Comparative genomics has provided valuable insights on plant peptidase inhibitor families and could explain the evolutionary reasons that lead to the current variable repertoire of peptidase inhibitors in specific plant clades., Ministerio de Educación y Ciencia, Ministerio de Economía y Competitividad, Comisión Europea FP7, Depto. de Bioquímica y Biología Molecular, Fac. de Ciencias Biológicas, TRUE, pub