Curated Optogenetic Publication Database

Abstract: BAR (Bin, Amphiphysin and Rvs) protein domains are responsible for the generation of membrane curvature and represent a critical mechanical component of cellular functions. Thus, BAR domains have great potential as components of membrane-remodeling tools for cell biologists. In this work, we describe the design and implementation of a family of versatile light-gated I-BAR (inverse-BAR) domain containing tools derived from the fusion of the A. thaliana Cryptochrome 2 photoreceptor and I-BAR protein domains ('CRY-BARs') with applications in the remodeling of membrane architectures and the control of cellular dynamics. By taking advantage of the intrinsic membrane binding propensity of the I-BAR domain, CRY-BARs can be used for spatial and temporal control of cellular processes that require induction of membrane protrusions. Using cell lines and primary neuron cultures, we demonstrate here that the CRY-BAR optogenetic tool evokes membrane dynamics changes associated with cellular activity. Moreover, we provide evidence that ezrin, an actin and PIP2 binding protein, acts as a relay between the plasma membrane and the actin cytoskeleton and therefore is an important mediator of switch function. Overall, we propose that CRY-BARs hold promise as a useful addition to the optogenetic toolkit to study membrane remodeling in live cells.

Abstract: The hallmarks of neurodegenerative diseases, including neural fibrils, reactive oxygen species (ROS), and cofilin-actin rods, present numerous challenges in the development of in vivo diagnostic tools. Biomarkers such as amyloid β (Aβ) fibrils and Tau tangles in Alzheimer's disease (AD) are accessible only via invasive cerebrospinal fluid assays, and ROS can be fleeting and challenging to monitor in vivo. Although remaining a challenge for in vivo detection, the protein-protein interactions underlying these disease-specific biomarkers present opportunities for the engineering of in vitro pathology-sensitive biosensors. These tools can be useful for investigating early-stage events in neurodegenerative diseases in both cellular and animal models and may lead to clinically useful reagents. Here, we report a light- and cellular stress-gated protein switch based on cofilin-actin rod formation, occurring in stressed neurons in the AD brain and following ischemia. By coupling the stress-sensitive cofilin-actin interaction with the light-responsive Cry2-CIB blue-light switch, referred to hereafter as the "CofActor," we accomplished both light- and energetic/oxidative stress-gated control of this interaction. Site-directed mutagenesis of both cofilin and actin revealed residues critical for sustaining or abrogating the light- and stress-gated response. Of note, the switch response varied, depending on whether cellular stress was generated via glycolytic inhibition or by both glycolytic inhibition and azide-induced ATP depletion. We also demonstrate light- and cellular stress-gated switch function in cultured hippocampal neurons. CofActor holds promise for the tracking of early-stage events in neurodegeneration and for investigating actin's interactions with other proteins during cellular stress.

Abstract: Elucidating temporal windows of signaling activity required for synaptic and behavioral plasticity is crucial for understanding molecular mechanisms underlying these phenomena. Here, we developed photoactivatable autocamtide inhibitory peptide 2 (paAIP2), a genetically encoded, light-inducible inhibitor of CaMKII activity. The photoactivation of paAIP2 in neurons for 1-2 min during the induction of LTP and structural LTP (sLTP) of dendritic spines inhibited these forms of plasticity in hippocampal slices of rodents. However, photoactivation ∼1 min after the induction did not affect them, suggesting that the initial 1 min of CaMKII activation is sufficient for inducing LTP and sLTP. Furthermore, the photoactivation of paAIP2 expressed in amygdalar neurons of mice during an inhibitory avoidance task revealed that CaMKII activity during, but not after, training is required for the memory formation. Thus, we demonstrated that paAIP2 is useful to elucidate the temporal window of CaMKII activation required for synaptic plasticity and learning.