Revisiting 2-Deoxyglucose Resistance in Neurospora crassa: Insights into Transcriptional Regulation of Hexose Transporters, Sugar Metabolism and Possible Modulation by Mating Type.

Abstract

Background: 2-Deoxyglucose (2DG) has been used as an antimetabolite since the 1950s, however, mechanisms associated with its toxicity continue to be evaluated. Studies on 2-DG resistance in Neurospora have identified and characterized some mutants, including rco-3 (NCU02582) and exo-1 (NCU09899). Compared to yeast what is known in terms of genes and mechanisms, our knowledge of this phenomenon in filamentous fungi is still sketchy, though growing. We felt that examining, various aspects of Carbon Catabolite Repression for a certain group of dgr mutants might provide us with some insights. Methods: In the present manuscript, we have employed physiological as well as transcriptional approaches to better understand various properties associated with dgr strains vis-à-vis other known hexose transporter knockouts ∆ hgt-1 (NCU10021), ∆ hgt-2 (NCU04963), ∆ glt-1 (NCU01633) as well as glucose sensors such as ∆rco-3 (NCU02582), the dgr mutants included in this study are dgr-L1 ‘a’, dgr-L1 ‘A’, dgr (BEX5) ‘a’ and dgr (BEX6) ‘a’. Results: We make the following observations regarding the phenotypes associated with the dgr strains (i.e. including dgr(L1), dgr (BEX5) and dgr (BEX6)). (i) In conditions involving both low and high glucose we see altered expression associated with hexose transporters (glt-1, hgt-1 and hgt-2) as well as xylose metabolism regulators (xlr-1) (ii) We observe differential response as measured in terms of growth with regards to mating type (iii) We also observe differential sensitivity with regards to pH of the dgr mutants (iv) Finally we also propose a rapid method by which Glucose uptake can be monitored microscopically using the fluorescent analogue 2-NBDG used to successfully validate uptake in the dgr mutants used in this study.