RESUMEN
Under dark anoxia, the unicellular green algae Chlamydomonas reinhardtii may produce hydrogen by means of its hydrogenase enzymes, in particular HYD1, using reductants derived from the degradation of intercellular carbon stores. Other enzymes belonging to the fermentative pathways compete for the same reductants. A complete understanding of the mechanisms determining the activation of one pathway rather than another will help us engineer Chlamydomonas for fermentative metabolite production, including hydrogen. We examined the expression pattern of the fermentative genes PDC3, LDH1, ADH2, PFL1, and PFR1 in response to day-night cycles, continuous light, continuous darkness, and low or high oxygen availability, which are all conditions that vary on a regular basis in Chlamydomonas' natural environment. We found that all genes except PFL1 show daily fluctuations in expression, and that PFR1 differentiated itself from the others in that it is clearly responsive to low oxygen, where as PDC3, LDH1, and ADH2 are primarily under diurnal regulation. Our results provide evidence that there exist at least three different regulatory mechanisms within the fermentative pathways and suggest that the fermentative pathways are not redundant but rather that availability of a variety of pathways allows for a differential metabolic response to different environmental conditions.
Asunto(s)
Chlamydomonas reinhardtii/enzimología , Chlamydomonas reinhardtii/genética , Fermentación/genética , Regulación de la Expresión Génica de las Plantas/genética , Secuencia de Bases , Ritmo Circadiano , Técnicas de Cultivo , ADN Complementario/análisis , Luz , Datos de Secuencia Molecular , Oxidorreductasas/genética , Oxidorreductasas/metabolismo , Oxígeno/metabolismo , ARN/análisis , TranscriptomaRESUMEN
⢠The unicellular green alga Chlamydomonas reinhardtii contains two iron (Fe)-hydrogenases which are responsible for hydrogen production under anoxia. In the present work the patterns of expression of alcohol dehydrogenase, a typical anaerobic gene in plants, of the hydrogenases genes (HYD1, HYD2) and of the genes responsible for their maturation (HYDEF, HYDG), were analysed. ⢠The expression patterns were analysed by real-time reverse-transcription polymerase chain reaction in Chlamydomonas cultures during the day-night cycle, as well as in response to oxygen availability. ⢠The results indicated that ADH1, HYD1, HYD2, HYDEF and HYDG were expressed following precise day-night fluctuations. ADH1 and HYD2 were modulated by the day-night cycle. Low oxygen plays an important role for the induction of HYD1, HYDEF and HYDG, while ADH1 and HYD2 expression was relatively insensitive to oxygen availability. ⢠The regulation of the anaerobic gene expression in Chlamydomonas is only partly explained by responses to anoxia. The cell cycle and light-dark cycles are equally important elements in the regulatory network modulating the anaerobic response in Chlamydomonas.
Asunto(s)
Alcohol Deshidrogenasa/genética , Chlamydomonas reinhardtii/enzimología , Chlamydomonas reinhardtii/genética , Ritmo Circadiano/genética , Hidrogenasas/genética , Adaptación Fisiológica/efectos de los fármacos , Adaptación Fisiológica/genética , Alcohol Deshidrogenasa/metabolismo , Anaerobiosis/efectos de los fármacos , Anaerobiosis/genética , Ciclo Celular/genética , Hipoxia de la Célula/genética , Supervivencia Celular/efectos de los fármacos , Chlamydomonas reinhardtii/citología , Chlamydomonas reinhardtii/efectos de los fármacos , Ritmo Circadiano/efectos de los fármacos , Medios de Cultivo/farmacología , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Genes de Plantas/genética , Hidrogenasas/metabolismo , Oxígeno/metabolismo , Oxígeno/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Solubilidad/efectos de los fármacos , Factores de TiempoRESUMEN
In this paper volatile organic compounds (VOCs) from durum wheat cultivars and landraces were analyzed using PTR-TOF-MS. The aim was to characterize the VOC's profile of the wholemeal flour and of the kernel to find out if any VOCs were specific to varieties and sample matrices. The VOC data is accompanied by SDS-PAGE analyses of the storage proteins (gliadins and glutenins). Statistical analyses was carried out both on the signals obtained by MS and on the protein profiles. The difference between the VOC profile of two cultivars or two preparations of the same sample - matrices, in this case kernel vs wholemeal flour - can be very subtle; the high resolution of PTR-TOF-MS - down to levels as low as pptv - made it possible to recognize these differences. The effects of grinding on the VOC profiles were analyzed using SIMPER and Tanglegram statistical methods. Our results show that it is possible describe samples using VOC profiles and protein data.