Discussion paper 8 Flashcards
What is the model organism of this paper?
Drosophila
What kind of synapse is being examined in this paper?
Synapse at the NMJ
What is the big-picture underlying question of this paper?
What mechanisms underlie synapse positioning and formation?
The cytoskeletal proteins being examined in this paper are…
Alpha and beta spectrin
What is the specific question being addressed by this paper?
What is the role of teneurins in regulating synapse formation and organization?
What are the most important proteins being examined by this paper with respect to synapse positioning and organization?
Teneurins
What 2 specific teneurins from drosophila are the focus of this paper?
Ten-a and ten-m
Teneurins were first identified in…
Vertebrates
Teneurins are enriched in…
The developing vertebrate brain
What question is being addressed in figure 1a?
What is the subcellular localization of teneurins at the NMJ?
What is horseradish peroxidase marking here?
Axon membrane
What are the key conclusions from this figure?
Ten-a localized in HRP zone – rough estimate for presynaptic membrane
Ten-m roughly in postsynaptic area
How do the authors address whether ten-a is expressed pre or postsynaptically?
Looking at the ten-a null mutant, compare to the ten-a neuron-specific knockdown and discover that ten-a is not expressed in muscle cells – must be expressed presynaptically (neurons only!)
Based on this image, is ten-m expressed pre or post synaptically?
Both
Describe the experimental approach here. What are M-IR and U-IR?
Using neuron and muscle specific knockdown approaches
M-IR = muscle antisense knockdown
U-IR = ubiquitous knockdown
What question do the authors address in this figure?
Where is the subcellular localization of ten-a
What is Fas2 a marker of?
Peri-active zone
What is Brp (bruchpilot) a marker of?
Active zone
What are the key conclusions from this figure?
Based on the partial overlap of Ten-a with Fas2 and Brp, Ten-a is localized in the junction between the active and peri-active zones
What is being shown in this figure and what conclusions can be made?
Ten-m is localized with Dlg and alpha-spectrin: due to this significant colocalization this confirms the finding that ten-m is associated with the postsynaptic membrane
What is the question being addressed by this figure?
Do ten-a and ten-m physically interact?
What is the experimental approach being used here?
Co-immunoprecipitation
Does this experiment show whether ten-a and ten-m directly interact?
No, they could be a part of a complex
What is the most important lane of this experiment and why?
Rightmost (specifically, top right) because it shows that ten-m and ten-a are interacting
What is the underlying question being addressed by this figure?
Are teneurins required for synaptogenesis?
What is being shown in this figure?
Muscle-specific knockdown of ten-m (b) and neural knockdown of ten-a (c) results in a deficit in formation of synaptic boutons compared to the wild type
What is the experimental procedure being performed in B?
Rna interference knockdown
What is happening in panel d?
Neural-specific rescue of ten-a results in wild-type phenotype (normal boutons)
What is being demonstrated by the rightmost bar in this graph?
That ten-a and ten-m must function in the same pathway for bouton genesis because rescue of ten-m has no effect on ten-a mutant – NOT REDUNDANT
In this figure, bouton (1) and (2) are being decreased by knockouts
Size and number
Based on the fact that in this graph there is not a complete reduction of synaptic boutons in this experiment….
There must be other molcules other than ten-a and ten-m which regulate bouton development
What are the other molecules which might mediate bouton development if not ten-a and ten-m?
Neuroligin/neurexin
What is the underlying question being addressed by this image?
Are teneurins required for correct synapse organization and structure?
What is the takeaway from this image?
Significant LACK of overlap between pre and post synaptic sites – deficit in structure
What question is being addressed here?
Are teneurin knockdowns associated with deficits in the structure of active zones?
What is being examined here?
Left: EPSP following motor nerve stimulation
Right: mEPSP (spontaneous)
Looking at response of motor nerve
What is the takeaway from this image?
Deficits in neurotransmitter release in both EPSP and mEPSP consistent with disruption of synapse development as seen earlier in the paper
What is the question being addressed in this figure?
Is there a physiological deficit in synapse function following teneurin knockdown (specifically ten-a)
What is the underlying question of this figure?
Are the NMJ synaptic defects correlated with cytoskeletal defects?
What is Futsch?
Labels microtubules
Dlg-1 is a scaffold protein which…
Adjusts the size of the post-synaptic surface
What are the most important findings here?
Grainy futsch (indicated by arrows in b and c) represents a lack of MT bundles compared to the control
Why does panel b represent a non cell autonomous defect?
Looking at muscle cells, but there are defects in the presynaptic cells too which means postsynaptic defects impact presynaptic cells
What are the most important findings of this image?
Mutating either teneurin results in a deficit of alpha spectrin postsynaptically, but does not impact Dlg postsynaptically
What is the question being addressed in this image?
Do alpha spectrin and teneurin-m physically interact?
What is the key takeaway from this image?
Alpha spectrin and ten-m physically interact
What is the question being addressed in this figure?
Are teneurins sufficient to specify synaptic target selection?
What is the experimental approach being used in this figure?
Use h94-gal4 line to overexpress ten-m in muscle 6, which normally does not express ten-m and see if this changes the connectivity of motor axon afferents (alters the ratio)
What is the assay in this figure?
Quantification of synaptic boutons in muscle 7/6
What is the key takeaway from this figure?
Overexpression of ten-m is sufficient to alter target selectivity
