Characterization of genetic and molecular tools for studying the endogenous expression ofLactate dehydrogenaseinDrosophila melanogaster

Abstract

Drosophila melanogaster larval development relies on a specialized metabolic state that utilizes carbohydrates and other dietary nutrients to promote rapid growth. One unique feature of the larval metabolic program is that Lactate Dehydrogenase (Ldh) activity is highly elevated during this growth phase when compared to other stages of the fly life cycle, indicating that Ldh serves a key role in promoting juvenile development. Previous studies of larval Ldh activity have largely focused on the function of this enzyme at the whole animal level, however, Ldh expression varies significantly among larval tissues, raising the question of how this enzyme promotes tissue-specific growth programs. Here we characterize two transgene reporters and an antibody that can be used to study Ldh expression in vivo. We find that all three tools produce similar Ldh expression patterns. Moreover, these reagents demonstrate that the larval Ldh expression pattern is complex, suggesting the purpose of this enzyme varies across cell types. Overall, our studies validate a series of genetic and molecular reagents that can be used to study glycolytic metabolism in the fly.

Glycolytic Disruption Triggers Interorgan Signaling to Nonautonomously RestrictDrosophilaLarval Growth

“…The upd3 mutant strain was obtained from the BDSC (RRID:BDSC_55728). The Ldh-GFP Genomic stock (RRID:BDSC_94704) used for colocalization analysis of Ldh and Gpdh1 was previously described (Bawa et al, 2020;Rai et al, 2024). The w [1118]; P{w[+mC]=10XStat92E-GFP}2 stock was obtained from the BDSC (RRID:BDSC_26198) and recombined with Ldh 16 to visualize STAT expression levels in the tissues of (Gpdh1 A10/B18 ) double mutant larvae.…”

“…Ldh protein expression was visualized using an anti-Ldh antibody (Boster bio DZ41222) as previously described (Rai et al, 2024). For larval tissues staining with anti-Gpdh1 antibody (Boster Bio DZ41223), the same protocol was used as for anti-Ldh staining, except that the anti-Gpdh1 antibody was diluted 1:100.…”

“…Intriguingly, while the loss of either single enzyme has minimal effect on overall larval growth, Gpdh1; Ldh double mutants experience a developmental arrest, suggesting a functional redundancy, where each enzyme partially compensates for the loss of the other (Li et al, 2019). However, neither Ldh nor Gpdh1 are ubiquitously expressed in larvae (Rechsteiner, 1970, Li et al, 2019, Rai et al, 2024, raising questions as to how these enzymes serve compensatory roles if not expressed in a strictly overlapping pattern. Here we address this question by examining the tissuespecific functions of both enzymes.…”