Welcome to Irini's WormWorld
Dense-core vesicle biogenesis
Neuromodulation is the process in which neurons and neuroendocrine cells secrete hormones, growth factors, and monoamines (e.g. dopamine, serotonin) to modulate the activity of a diverse population of cells. Dysfunction of these secretory pathways can lead to numerous metabolic and neurological diseases, ranging from diabetes to mental disorders.
The regulated release of neuromodulators is mediated by a unique type of cellular vesicle, the dense-core vesicle (DCV). DCVs originate from the trans-Golgi, but it is unclear how they are formed and acquire their specific cargos. To identify proteins that act in DCV biogenesis, we performed a forward genetic screen in C. elegans for mutants defective in DCV function. We identified mutations in the small GTPase RAB-2, the RAB-2 effectors RUND-1/RUNDC1 (a RUN domain protein) and CCCP-1/CCDC186 (a coiled-coil protein), the endosome-associated recycling protein (EARP) complex and its interactor EIPR-1. Genetic epistasis data indicate that RAB-2/CCCP1/RUND1 and EIPR-1/EARP function in a common pathway (Fig. 1.
Using insulin-secreting insulinoma cells we found that in the absence of EIPR1 or CCDC186 cells secrete less insulin while mature DCV cargoes accumulate near the trans-Golgi network and are not retained in mature DCVs in the cell periphery (Fig. 2). Also, EIPR1 and CCDC186 KO lines retain carboxypeptidase D in mature DCVs, a cargo that in wild type cells is found only in immature DCVs. Our working hypothesis is that CCDC186 and EARP act together to control the post-Golgi retention and removal of cargos from mature DCVs (Cattin et al, 2019).
In addition we showed that EIPR1 is a strong interactor of the EARP complex and that it is required for the stability of the EARP complex subunits, for the localization of EARP, and its association with membranes. EARP is localized to two distinct compartments related to its function: an endosomal compartment and a DCV biogenesis-related compartment. Our working hypothesis is that EIPR1 functions together with EARP to control both endocytic recycling and DCV maturation (Fig. 1.
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Fig. 1
Fig. 2
Dopaminergic signaling negatively regulates
NALCN/NCA ion channels
The NALCN/NCA ion channel is a cation channel related to voltage-gated sodium and calcium channels. NALCN has been reported to be a sodium leak channel, which regulates the resting membrane potential and excitability of invertebrate and vertebrate neurons, but its precise cellular role and regulation are unclear.
The C. elegans orthologs of NALCN, NCA-1 and NCA-2, act in premotor interneurons to regulate motor circuit activity that sustains locomotion. Recently, we found that NCA-1 and NCA-2 are activated by a signal transduction pathway acting downstream of the heterotrimeric G protein Gq and the small GTPase Rho (Topalidou et al, 2017a). Through a forward genetic screen, I identified the GPCR kinase GRK-2 as a new player affecting signaling through the Gq-Rho-NCA pathway. I showed that GRK-2 acts on the D2-like dopamine receptor DOP-3 to inhibit Go signaling and positively modulate NCA-1 and NCA-2 activity (Fig. 3. Topalidou et al, 2017b). This pathway is conserved in mammals, as a recent study following up on our work found that dopamine acts through D2 receptors and Gi/o to inhibit NALCN in the mouse brain.
To advance my genetic observations I collaborated with the Bai lab (Fred Hutch) and used an array of behavioral, electrophysiological, biochemical, and imaging approaches to determine the molecular mechanisms that connect dopaminergic signaling to the NCA activity. Our preliminary data show that GRK-2 and DOP-3 act in opposite way in premotor interneurons to modulate the activity of the NCA channel. Our working hypothesis is that dopamine modulates the activity of the NCA channels through GRK-2.
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Sensory neurons regulate C. elegans locomotion quiescence through GRK-2 signaling
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Fig. 3
C. elegans goes through periods of behavioral quiescence during larval molts (lethargus) and locomotion quiescence as adults. Previous work has shown that both lethargus and adult locomotion quiescence is reduced in mutants lacking the neuropeptide receptor NPR-1. In recent experiments I found that mutants in the GPCR kinase GRK-2 show enhanced locomotion quiescence and that npr-1 mutants suppress this phenotype. To verify my observations I collaborated with the Raizen lab and found that GRK-2 regulates locomotion quiescence in a manner largely independent of the transcription factors APTF-1 and CEH-17 that regulate lethargus quiescence and stress-induced quiescence, respectively. Using neuron-specific rescuing experiments I showed that GRK-2 acts in multiple ciliated sensory neurons to mediate this behavior. My working hypothesis is that sensory neurons regulate locomotion quiescence through GRK-2 signaling (Fig. 4).
Fig. 4
EDUCATION
1999 - 2003
University of Crete, Heraklion, Greece
PhD Program in Molecular Biology & Biomedicine, Department of Biology
1997 - 1999
University of Crete, Heraklion, Greece
MSc Program in Molecular Biology & Biomedicine, Department of Biology
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1992 - 1996
National & Kapodistrian university of Athens, Athens, Greece
BSc Biology, Department of Biology
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2019-present
Senior Scientist
University of Washington, Biochemistry Department, Seattle, WA
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2018-2019
Research Scientist IV
University of Washington, Biochemistry Department, Seattle, WA
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2017-2018
Research Scientist III
University of Washington, Biochemistry Department, Seattle, WA
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2012-2017
Acting Instructor
University of Washington, Biochemistry Department, Seattle, WA
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2009-2012
Research Scientist Associate
Columbia University, Dept. of Biological Sciences, New York, NY
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2004-2009
Postdoctoral Fellow
Columbia University, Dept. of Biological Sciences, New York, NY