Passos iniciais do metabolismo de isoprenoides em angiospermas: genômica evolutiva e análises moleculares em Coffea spp

Abstract

Os isoprenoides compõem uma classe de metabólitos especializados em plantas, com funções biológicas, ecológicas e fisiológicas. As plantas sintetizam milhares de compostos derivados dos isoprenos, com função associada a atração de polinizadores, proteção contra pragas, desenvolvimento e crescimento das plantas. Além disso, eles são muito utilizados pelas indústrias farmacêutica e de cosméticos. Existem duas vias metabólicas capazes de produzir isoprenoides em plantas: a via citoplasmática do mevalonato (MVA) e a via plastidial do metileritritol fosfato (MEP). Os objetivos do trabalho foram: i) identificar os genes iniciais da via de biossíntese de isoprenoides em Coffea spp.; ii) avaliar se o elicitor ácido hexanoico (Hx) é capaz de modular o perfil transcricional dos genes da via MVA e MEP em C. arabica; iii) analisar a evolução dos genes chave da via MVA (HMGR e MVK) e MEP (DXS e DXR). Plantas de C. arabica cv. Catuaí Vermelho (CaCV) e cv. Obatã (CaOB) foram tratadas com aplicação de ácido hexanóico (Hx - 0,55 mM) em solução nutritiva aerada, e após 48 h foram coletadas folhas e raízes para extração de RNA e análises de perfil transcricional via RNA-seq e RT-qPCR. Foram identificados mais de uma cópia para cada gene da via MVA (HMGS, HMGR, MVK, PMK, MVD) e MEP (DXS, DXR, CMK, MDS, HDS, HDR) em C. arabica. C. eugenioides apresentou mais de uma cópia para alguns genes da via MVA (AACT, HMGR, PMK, MVD) e da via MEP (DXS e HDR). Em C. canephora, os genes da MVA (AACT, HMGS, MVK, PMK, MVD) e alguns da MEP (DXR, MCT, CMK, MDS, HDS) foram de cópia única. Os genes AACT, HMGS, HMGR, MVK, PMK, MVD, DXS, DXR, CMK e MDS foram induzidos em folhas tratadas com Hx em CaCV. Em CaOB, os genes MVK, DXS, DXR, CMK, MDS e HDS foram induzidos em folhas tratados com Hx. Os genes AACT, HMGS, HMGR, MVK, PMK, DXS e DXR foram reprimidos em raízes de CaCV e os genes AACT, HMGS, HMGR, MVK, MVD, DXS, DXR e MDS reprimidos em raízes de CaOB tratadas com Hx. Os nossos resultados indicam que o Hx é capaz de modular a expressão dos genes das vias MVA e MEP, em folhas e raízes de café de maneira genótipo e tecido-específica, evidenciando o poder reprogramador de Hx no metabolismo de isoprenoides.

Isopropenoids comprise a class of specialized metabolites from plants comprising important biological, ecological and physiological functions. Plants synthesize thousands of isoprene compounds, which are related to pollinators attraction, antiherbivory properties, plant growth and development. In addition, they are widely used by the pharmaceutical and cosmetic industries. There are two metabolic pathways responsible for isoprenoid biosynthesis in plants: the cytoplasmic mevalonate pathway (MVA) and the plastidic metileritritol phosphate pathway (MEP). In this study our main goals were: i) to identify genes related to the initial steps of isoprenoid biosynthesis in Coffea spp.; ii) evaluate if hexanoic acid (Hx) can transcriptionally modulate genes of MVA and MEP pathway in C. arabica; iii) analyze the evolution of selected MVA (HMGR and MVK) and MEP (DXS and DXR) genes. C. arabica cv. Catuaí Vermelho (CaCV) and cv. Obatã (CaOB) plants were treated with exogenous application of hexanoic acid (Hx - 0.55 mM) in an aerated nutrient solution, and after 48 h, leaves and roots were collected for RNA extraction and transcriptional analyses using RNAseq and RT-qPCR. More than one copy were observed for several MVA pathway genes (HMGS, HMGR, MVK, PMK, MVD) and MEP pathway genes (DXS, DXR, CMK, MDS, HDS, HDR) in C. arabica. C. eugenioides presented more than one copy of some genes from MVA (AACT, HMGR, PMK, MVD) and MEP (DXS, HDR) pathways. In C. canephora, some genes from MVA (AACT, HMGS, MVK, PMK, MVD) and MEP (DXR, MCT, CMK, MDS, HDS) pathways were classified as single copy. AACT, HMGS, HMGR, MVK, PMK, MVD, DXS, DXR, CMK and MDS genes were induced in leaves under Hx-treatment in CaCV. In CaOB, MVK, DXS, DXR, CMK, MDS and HDS genes were induced in leaves by Hx exogenous application. AACT, HMGS, HMGR, MVK, PMK, DXS and DXR genes were repressed in CaCV roots and AACT, HMGS, HMGR, MVK, MVD, DXS, DXR e MDS repressed in CaOB roots under Hx treatment. Our results indicated that Hx can modulate the expression of genes involved in MVA and MEP pathways in coffee in a genotype and tissue-specific way, emphasizing that Hx is able to reprogram isoprenoid metabolism.