The glycolytic pathway is a catabolic route that ends
The glycolytic pathway is a catabolic route that ends with the synthesis of pyruvate (Pyr). In Arabidopsis embryos, the conversion of phosphoenolpyruvate (PEP) to Pyr, catalyzed by pyruvate kinase (PK), is mainly located in the plastids , . The preceding reaction is catalyzed by enolase (ENO) which insures the Mg2+-dependent conversion of 2-phosphoglycerate (2-PGA) to PEP. Plant ENOs are encoded by a small gene family , . Previous studies have provided evidence that Arabidopsis ENO isoforms are present in both the plastid and the cytosol , . Furthermore, in Arabidopsis, cytosolic glycolysis is thought to provide PEP necessary to support plastidial oil production . Although, recently, the participation of the plastidial glycolysis has been claimed as a potential contributor to the process .
Previously, we have shown that in sunflower embryos crude extracts ENO activity increases during development in conjunction with oil deposition, and that most of this ENO activity is associated to the cytosol . These data could indicate that, in developing sunflower seeds, oil accumulation is mainly supported by PEP generated in the cytosol. However, further characterization of the different ENOs expressed in the embryos is required in order to better understand the participation of cytosolic and plastidial glycolysis during oil synthesis.
In this study, we have identified and characterized three sunflower ENO isoforms. We surveyed the 1568 of ENO genes and the subcellular localization of the corresponding protein products. Recombinant ENO proteins were expressed in Escherichia coli in order to determine their kinetic properties. Similarly, to the situation observed in A. thaliana, we found that one the three ENO proteins did not have catalytic activity. Site directed mutagenesis was used to recover low levels of activity for this protein. These data extend our understanding of the contribution of ENO isoforms to the conversion of 2-PGA to PEP in the cytosolic and plastidial compartments of sunflower seeds during fatty acid synthesis. Our results are discussed in relation to a model that recapitulates current knowledge on the importance of the glycolytic pathway in sunflower seed filling.
Materials and methods
Discussion In developing seeds, glycolysis is the paramount metabolic pathway in which hexoses are converted to Pyr that is subsequently being used as a source of carbon for fatty acids biosynthesis within plastids. PEP, the product of ENO activity, is the immediate precursor of Pyr and therefore plays a key biochemical function during lipid deposition. ENO is a ubiquitous enzyme that is present in the plant cytosolic and plastidial compartments. The present study was undertaken to characterize the ENO gene family in sunflower in relation to seed development and lipid reserve accumulation.
Acknowledgements This work was supported by the “Ministerio de Economía y Competitividad” and FEDER (AGL2014-53537-R). JR is supported by a Discovery Grant (RGPIN 227271) from the National Science and Engineering Research Council of Canada. The authors would also like to thank the Hauts-de-France Region and the European Regional Development Fund (ERDF) 2014/2020 for the funding of this work.
Introduction Tuberculosis (TB) is a global disease caused by Mycobacterium tuberculosis (Mtb), resulting in serious disease burden and death of human population in millions. This alarming scenario is the case when approximately one third of infections in humans remain asymptomatic or latent . Mtb is a facultative intracellular human pathogen which has the ability to grow extracellularly, invade host tissues and spread systemically leading to a debilitating disease. Upon reactivation of latent infection, due to factors such as aging or immunosuppression, the bacilli can induce an active spread of the disease dependent on host matrix metalloproteinases (MMPs) leading to tissue degeneration . MMPs are endopeptidases capable of degrading components of the extracellular matrix (ECM) like collagen .