Spectrin Tetramer Formation Is Not Required for Viable Development in Drosophila

Khanna, Mansi R.; Mattie, Floyd J.; Browder, Kristen C.; Radyk, Megan D.; Crilly, Stephanie E; Bakerink, Katelyn J.; Harper, Sandra L.; Speicher, David W.; Thomas, Graham H.

doi:10.1074/jbc.m114.615427

Cited by 15 publications

(15 citation statements)

References 56 publications

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“…Claire H. Thomas (Khanna et al, 2015) N/A N/A and HRP (blue). Expression of cytoplasmic GFP1-10 in -Spec spGFP11 animals triggered compartment specific GFP reconstitution.…”

Section: Uas-myc-α-specmentioning

confidence: 99%

A presynaptic spectrin network controls active zone assembly and neurotransmitter release

Wang

Friend

Vicidomini

et al. 2019

Preprint

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We have previously reported that Drosophila Tenectin (Tnc) recruits PS2/PS integrin to ensure structural and functional integrity at larval NMJs (Wang et al., 2018). In muscles, Tnc/integrin engages the spectrin network to regulate the size and architecture of synaptic boutons. In neurons, Tnc/integrin controls neurotransmitter release. Here we show that presynaptic Tnc/integrin modulates the synaptic accumulation of key active zone components, including the Ca 2+ channel Cac and the active zone scaffold Brp. Presynaptic α-Spectrin appears to be both required and sufficient for the recruitment of Cac and Brp. We visualized the endogenous α-Spectrin and found that Tnc controls spectrin recruitment at synaptic terminals. Thus, Tnc/integrin anchors the presynaptic spectrin network and ensures the proper assembly and function of the active zones. Since pre-and postsynaptic Tnc/integrin limit each other, we hypothesize that this pathway links dynamic changes within the synaptic cleft to changes in synaptic structure and function. The synaptic extracellular matrix (ECM) contains various synaptic organizers and diffusible molecules that establish and modify synaptic connections during development. ECM receptors, including integrins, transduce these signals by sensing the changes in the synaptic environment and orchestrating the appropriate changes in synapse structure and function (Chavis and Westbrook, 2001;Huang et al., 2006). Disruption of integrins impair synapse development and maturation, neurotransmission, and long-term synaptic plasticity (reviewed in (Park and Goda, 2016)). Integrins together with their ECM ligands form complexes with cytoskeletal proteins and active zone (AZ) components (Carlson et al., 2010;Nishimune et al., 2004;Sunderland et al., 2000). However, it remains unclear how these interactions could influence neurotransmitter release.Spectrin, a principal component of the membrane skeleton, is critical for synaptic transmission (Featherstone et al., 2001;Goodman, 1999). Spectrin subunits function as docking platforms for membrane channels, neurotransmitter receptors and adhesion molecules (reviewed in (Machnicka et al., 2014)). For example, spectrins associate with presynaptic Ca 2+ channels in synaptosome lysates (Carlson et al., 2010;Khanna et al., 2007a;Khanna et al., 2007b). Also, a spectrin-containing presynaptic web enables AZ function in the rodent CNS (Phillips et al., 2001). Recent super-resolution fluorescence microscopy uncovered periodic actin/spectrin lattices along axons, dendrites and spines in a broad range of neuronal cell types (He et al., 2016;Qu et al., 2017;Xu et al., 2013); however, at presynaptic and postsynaptic sites, these patterns were absent (Sidenstein et al., 2016).We have previously found that Drosophila Tenectin (Tnc), an integrin ligand that accumulates in the synaptic cleft at the larval NMJ, modulates synapse development and function (Wang et al., 2018). Tnc recruits αPS2/βPS integrin and forms cis active complexes with distinct pre-and post-synaptic functions....

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“…Claire H. Thomas (Khanna et al, 2015) N/A N/A and HRP (blue). Expression of cytoplasmic GFP1-10 in -Spec spGFP11 animals triggered compartment specific GFP reconstitution.…”

Section: Uas-myc-α-specmentioning

confidence: 99%

A presynaptic spectrin network controls active zone assembly and neurotransmitter release

Wang

Friend

Vicidomini

et al. 2019

Preprint

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“…Second, although spectrin is essential to maintain the integrity of axons when they experience large rapid stresses and strains, it does not play a central role in the process of wiring the nervous system (61). In particular, animals null for a-spectrin or expressing mutated versions of a-spectrin, which inhibit tetramer formation, have very mild defects in the organization of the nervous system (62,63).…”

Section: Computational Modelmentioning

confidence: 99%

Modeling the Axon as an Active Partner with the Growth Cone in Axonal Elongation

Rooij

Kuhl

Miller

2018

Biophysical Journal

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Forces generated by the growth cone are vital for the proper development of the axon and thus brain function. Although recent experiments show that forces are generated along the axon, it is unknown whether the axon plays a direct role in controlling growth cone advance. Here, we use analytic and finite element modeling of microtubule dynamics and the activity of the molecular motors myosin and dynein to investigate mechanical force balance along the length of the axon and its effects on axonal outgrowth. Our modeling indicates that the paradoxical effects of stabilizing microtubules and the consequences of microtubule disassembly on axonal outgrowth can be explained by changes in the passive and active mechanical properties of axons. Our findings suggest that a full understanding of growth cone motility requires a consideration of the mechanical contributions of the axon. Our study not only has potential applications during neurodevelopment but might also help identify strategies to manipulate and promote axonal regrowth to treat neurodegeneration.

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“…In addition, Penn State Hershey College of Medicine, as well as some external research labs and universities, have used the facility. A number of exciting publications have ensued as a result of the capability [Khanna et al , 2015; Warui et al , 2015; Bastidas et al , 2013; Liu et al , 2013; Toroney et al , 2012; Patel et al , 2012; Mukherjee et al , 2011]. The speed of the Automated ITC and DSC combined with access to additional supporting instrumentation helps streamline research data collection considerably.…”

Section: Science Facilitatedmentioning

confidence: 99%

“…Binding of menadione and quinone with proteins of the photosystem I reaction center were studied. Three different buffer concentrations were used in the preliminary phase to quickly determine best buffer conditions.The Thomas lab from the department of Biochemistry and Molecular Biology used the Auto-iTC200 to quantify the oligomerization-interaction between the tetramerization domains of α, β and β [heavy] spectrins and several variants from the fruit fly genus Drosophila [Khanna et al , 2015]. …”

Section: Science Facilitatedmentioning

confidence: 99%

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A High-Throughput Biological Calorimetry Core

Yennawar

Fecko

Showalter

et al. 2016

Methods in Enzymology

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Many labs have conventional calorimeters where denaturation and binding experiments are setup and run one at a time. While these systems are highly informative to biopolymer folding and ligand interaction, they require considerable manual intervention for cleaning and setup. As such, the throughput for such setups is limited typically to a few runs a day. With a large number of experimental parameters to explore including different buffers, macromolecule concentrations, temperatures, ligands, mutants, controls, replicates, and instrument tests, the need for high-throughput automated calorimeters is on the rise. Lower sample volume requirements and reduced user intervention time compared to the manual instruments have improved turnover of calorimetry experiments in a high-throughput format where 25 or more runs can be conducted per day. The cost and efforts to maintain high-throughput equipment typically demands that these instruments be housed in a multiuser core facility. We describe here the steps taken to successfully start and run an automated biological calorimetry facility at Pennsylvania State University. Scientists from various departments at Penn State including Chemistry, Biochemistry and Molecular Biology, Bioengineering, Biology, Food Science, and Chemical Engineering are benefiting from this core facility. Samples studied include proteins, nucleic acids, sugars, lipids, synthetic polymers, small molecules, natural products, and virus capsids. This facility has led to higher throughput of data, which has been leveraged into grant support, attracting new faculty hire and has led to some exciting publications.

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Spectrin Tetramer Formation Is Not Required for Viable Development in Drosophila

Cited by 15 publications

References 56 publications

A presynaptic spectrin network controls active zone assembly and neurotransmitter release

A presynaptic spectrin network controls active zone assembly and neurotransmitter release

Modeling the Axon as an Active Partner with the Growth Cone in Axonal Elongation

A High-Throughput Biological Calorimetry Core

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