Biomedical subjects

E M Skoulakis

Publications and source records attributed to E M Skoulakis.

7 recordsLinked to original sources

PubMed2001

Conditional rescue of olfactory learning and memory defects in mutants of the 14-3-3zeta gene leonardo.

Members of the ubiquitous 14-3-3 family of proteins are abundantly expressed in metazoan neurons. The Drosophila 14-3-3zeta gene leonardo is preferentially expressed in adult mushroom bodies, centers of insect learning and memory. Mutants exhibit defects in olfactory learning and memory and physiological neuroplasticity at the neuromuscular junction. Because strong mutations in this gene are lethal, we investigated the nature of the defects that precipitate the learning and memory deficit and the role of the two protein isoforms encoded by leonardo in these processes. We find that the behavioral deficit in the mutants is not caused by aberrant development, LEONARDO protein is acutely required for learning and memory, and both protein isoforms can function equivalently in embryonic development and behavioral neuroplasticity.

14-3-3 Proteins↗

PubMed1998

14-3-3 proteins in neuronal development and function.

The 14-3-3 proteins are small, cytosolic, evolutionarily conserved proteins expressed abundantly in the nervous system. Although they were discovered more than 30 yr ago, their function in the nervous system has remained enigmatic. Several recent studies have helped to clarify their biological function. Crystallographic investigations have revealed that 14-3-3 proteins exist as dimers and that they contain a specific region for binding to other proteins. The interacting proteins, in turn, contain a 14-3-3 binding motif; proteins that interact with 14-3-3 dimers include PKC and Raf, protein kinases with critical roles in neuronal signaling. These proteins are capable of activating Raf in vitro, and this role has been verified by in vivo studies in Drosophila. Most interestingly, mutations in the Drosophila 14-3-3 genes disrupt neuronal differentiation, synaptic plasticity, and behavioral plasticity, establishing a role for these proteins in the development and function of the nervous system.

14-3-3 Proteins↗

PubMed1997

Leonardo, a Drosophila 14-3-3 protein involved in learning, regulates presynaptic function.

The leonardo gene encodes a conserved member of the 14-3-3 protein family, which plays a role in Drosophila learning. Immunological localization of the protein shows that it is expressed at synaptic connections and enriched in presynaptic boutons of the neuromuscular junction (NMJ). Null leonardo mutants die as mature embryos. Electrophysiological assays of the mutant NMJ demonstrate that basal synaptic transmission is reduced by 30% and that transmission amplitude, fidelity, and fatigue resistance properties are reduced at elevated stimulation frequencies and in low external [Ca2+]. Moreover, transmission augmentation and post-tetanic potentiation (PTP) are disrupted in the mutant. These results suggest that Leonardo plays a role in the regulation of synaptic vesicle dynamics, a function which may underlie synaptic modulation properties enabling learning.

14-3-3 Proteins↗

PubMed1997

The Drosophila 14-3-3 protein Leonardo enhances Torso signaling through D-Raf in a Ras 1-dependent manner.

14-3-3 proteins have been shown to interact with Raf-1 and cause its activation when overexpressed. However, their precise role in Raf-1 activation is still enigmatic, as they are ubiquitously present in cells and found to associate with Raf-1 in vivo regardless of its activation state. We have analyzed the function of the Drosophila 14-3-3 gene leonardo (leo) in the Torso (Tor) receptor tyrosine kinase (RTK) pathway. In the syncytial blastoderm embryo, activation of Tor triggers the Ras/Raf/MEK pathway that controls the transcription of tailless (tll). We find that, in the absence of Tor, overexpression of leo is sufficient to activate tll expression. The effect of leo requires D-Raf and Ras1 activities but not KSR or DOS, two recently identified essential components of Drosophila RTK signaling pathways. Tor signaling is impaired in embryos derived from females lacking maternal expression of leo. We propose that binding to 14-3-3 by Raf is necessary but not sufficient for the activation of Raf and that overexpressed Drosophila 14-3-3 requires Ras1 to activate D-Raf.

14-3-3 Proteins↗

PubMed1996

Olfactory learning deficits in mutants for leonardo, a Drosophila gene encoding a 14-3-3 protein.

Studies of Drosophila and other insects have indicated an essential role for the mushroom bodies in learning and memory. The leonardo gene encodes a Drosophila protein highly homologous to the vertebrate 14-3-3zeta isoform, a protein well studied for biochemical roles but without a well established biological function. The gene is expressed abundantly and preferentially in mushroom body neurons. Mutant alleles that reduce LEONARDO protein levels in the mushroom bodies significantly decrease the capacity for olfactory learning, but do not affect sensory modalities or brain neuroanatomy that are requisite for conditioning. These results establish a biological role for 14-3-3 proteins in mushroom body-mediated learning and memory processes, and suggest that proteins known to interact with them, such as RAF-1 or other protein kinases, may also have this biological function.

14-3-3 Proteins↗

PubMed1993

Preferential expression in mushroom bodies of the catalytic subunit of protein kinase A and its role in learning and memory.

Involvement of the cAMP cascade in Drosophila learning and memory is suggested by the aberrant behavioral phenotypes of the mutants dunce (cAMP phosphodiesterase) and rutabaga (adenylyl cyclase). Line DCO581, isolated via an enhancer detector screen for genes preferentially expressed in the mushroom bodies, contains a transposon in the first exon of the catalytic subunit gene (DCO) of protein kinase A (PKA). RNA in situ hybridization and immunohistochemistry show that DCO is preferentially expressed in the mushroom bodies. The DCO581 insertion and an independently isolated hypomorphic allele (DCOB10) each produce homozygous lethality and a 40% decrease in PKA activity in heterozygotes. This decrease has mild effects on learning but no effect on memory. However, the 80% reduction in activity obtained by constructing heteroallelic yet viable DCO581/DCOB10 animals results in a dramatic learning and memory deficit. These results suggest that PKA plays a crucial role in the cAMP cascade in mushroom bodies to mediate learning and memory processes.

Animals↗

PubMedDate not supplied

Tripartite mushroom body architecture revealed by antigenic markers.

We have explored the organization of the axonal lobes in Drosophila mushroom bodies by using a panel of immunohistochemical markers. These markers consist of antibodies to eight proteins expressed preferentially in the mushroom bodies: DAMB, DCO, DRK, FASII, LEO, OAMB, PKA RII, and RUT. Previous to this work, four axonal lobes, two projecting dorsally (alpha and alpha') and two medially (beta and gamma), had been described in Drosophila mushroom bodies. However, our analysis of immunohistochemically stained frontal and sagittal sections of the brain revealed three medially projecting lobes. The newly distinguished lobe, which we term beta', lies along the dorsal surface of beta, just posterior to gamma. In addition to resolving a fifth lobe, our studies revealed that there are specific lobe sets defined by equivalent marker expression levels. These sets are (1) the alpha and beta lobes, (2) the alpha' and beta' lobes, and (3) the gamma lobe and heel (a lateral projection formed by a hairpin turn of some of the peduncle fibers). All of the markers we have examined are consistent with these three sets. Previous Golgi studies demonstrate that each mushroom body cell projects one axon that branches into a dorsal lobe and a medial lobe, or one unbranched axon that projects medially. Taken together with the lobe sets listed above, we propose that there are three major projection configurations of mushroom body cell axons: (1) one branch in the alpha and one in the beta lobe, (2) one branch in the alpha' and one in the beta' lobe, and (3) one unbranched axon projecting to the heel and the gamma lobe. The fact that these neuron types exhibit differential expression levels of a number of mushroom body genes suggests that they may have corresponding functional differences. These functions may be conserved in the larvae, as several of these genes were expressed in larval and embryonic mushroom bodies as well. The basic mushroom body structure, including the denritic calyx, peduncle, and lobes, was already visible by the late stages of embryogenesis. With new insights into mushroom body organization, and the characterization of markers for developing mushroom bodies, we are beginning to understand how these structures form and function.

Animals↗