The glycolytic pathway is the best understood in biology. For this pathway, it is proposed that individual enzyme levels are matched to the maximum metabolic flux demands of a particular cell type . An extreme example of this essential matching is insect flight, where the highest mass-specific oxygen consumption rates among animals are observed. In the flight muscle, high enzyme levels are needed for the rapid turnover of ATP associated with the 100-fold or greater increase in oxygen consumption over resting states . Wing beat frequency (WBF) is closely correlated with oxygen consumption and directly reflects the rate of ATP hydrolysis and glycolytic flux. This tight connection between pathway flux and ATP turnover makes the measurement of WBF an ideal system for studying metabolic flux control.
We used P-element excision-derived knockouts in Drosophila melanogaster to experimentally lower the in vivo activity levels of seven enzymes in this pathway and examine individual enzyme influence on tethered flight performance measured as wing beat frequency (WBF) under high speed video and ability to sustain free flight (Flowers et al. 2006). Our results find the classic regulatory enzyme, glycogen phosphorylase, is nearest to capacity or saturation, while other enzymes, especially those termed near-equilibrium, possess excess capacity. These latter enzymes show extreme genetic dominance and low flux control that extends down to markedly reduced enzyme levels. The genetic results are generally consistent with physiological studies using in vitro estimates that predict near-equilibrium enzymes carry excess capacity. The possibility is that this excess capacity is exploited under demand states different from the experimental test conditions.
I have an NSF-sponsored three year project (2009-2012) to continue these studies. This study will continue to study metabolic flight performance. examines the rate-limiting steps involved in pathway control (if any), and determines if there are differences in degree of saturation or excess capacity for different steps. These experiments test if this activity is drawn upon under conditions (temperature and load demands) requiring reserve capacity (formally these are genotype-by-environmental interactions). It will also explore the effects of gene-gene interactions (epistasis) on flight performance, and determines (1) if dominance can be modified by genetic background change, and (2) if dominance modification depends on the nature of mechanistic interrelationships between enzyme steps.
In insects, the phenotype most commonly associated with adaptation to temperate environments is the expression of a diapause syndrome. As with other Drosophila species, D. melanogaster overwinters as an adult, resulting in temporal population continuity. There is a phenotype in Drosophila melanogaster females that is called a reproductive diapause. Females, if placed upon emergence on a short day length and low temperature, show a response where egg production stops and developing eggs are absorbed. This trait is genetically variable in natural populations, and we have data showing that this trait is geographically variable with northern populations showing a much higher frequency of induction (Schmidt et al. 2005). The genetic variance for diapause has significant pleiotropic effects on a variety of other traits including lifespan, age-specific mortality rates, fecundity profiles, multiple forms of stress tolerance, lipid content, and development time. In natural populations of D. melanogaster in eastern North America, diapause genotype accounts for a significant amount of the observed variance for life histories that exhibit latitudinal clines (Schmidt and Paaby 2008).
have mapped the genetic variation in this trait to a specific gene (the couch
potato gene) and a single amino acid polymorphism that shows a parallel
geographic cline with diapause (Schmidt et al. 2008). I have a joint NSF-sponsored
three year grant (2009-2012) with Paul Schmidt
at the University of Pennsylvania to
further study cpo and its functional role in the establishment of diapause. The
project will examine the functional significance and adaptive dynamics of the cpo462Ile/Lys
polymorphism in natural populations. It will explore the global molecular
population genetics of the cpo
gene region and extend our analyses to ancestral African populations as well as
derived populations on other continents. We plan to identify modifiers of
diapause outside the third chromosome and to examine the interaction between cpo
and pathways known to affect life histories in Drosophila. To perform an initial
examination of the mechanism by which cpo
determines diapause and associated life histories.
challenge of balancing fluctuating nutrient inputs with the partitioning of
demand and storage is an essential feature of energy homeostasis that is common
to all life. Under nutrient excess, calories are stored and under nutrient
limitation this storage is drawn down. In natural populations this partitioning
is adjusted against demands of reproduction that trade off against lifespan. At
the genome level, transcriptional response to signals of nutrient level change
involves the activation and repression of major pathways associated with glucose
oxidation/fatty acid synthesis and fatty acid oxidation. These responses are the
outcome of signaling pathways in cells
such as the pancreas, fat body, and neurosecretory cells.
Evolution has led to a fine tuning of these signaling responses, and failure to
properly maintain energy homeostasis has significant impact on fitness.
common statement is that “nutrient sensors” or “sensory neurons”
initiate these signaling processes. These specialized cells sense the energy
state via the levels of metabolites. Since
the downstream signaling systems appear highly conserved in metazoans, we
propose that these upstream sensors are conserved as well. However, other than
the insulin secretion in mammals, very little is known in any model
species about the nutrient or metabolic energy-state sensors residing at the
very top that initiate signaling, or
how genetic variation in these sensors is involved in adaptation to
This project in collaboration with the lab of John True examines the extent to which nutrient signaling mechanisms are conserved between flies and mammals and examine Drosophila as a suitable model for understanding these mechanisms.