Contains a large deletion internal to the P{PZ}Hsc70-403550 insertion.
Hsc70-4Δ19/Df(3R)BSC471 larvae show a significant decrease in endosomal microautophagy at the neuromuscular junction compared to wild-type controls. Macroautophagy, synaptic vesicle endocytosis and proteasomal activity are not significantly different from wild type. Ultrastructurally, the boutons show no significant difference to wild type.
The mean excitatory junctional current (EJC) amplitude at the neuromuscular junction is significantly lower than wild type in Hsc70-4Δ19/Df(3R)BSC471 larvae.
Mutant neuromuscular junctions show reliable homeostatic compensation (increase in quantal content) after treatment with philanthotoxin-433 for 10 minutes.
Homozygous larvae show sporadic but reduced muscle contractions. The nerve-evoked excitatory junctional potential (EJP) amplitudes in homozygous mutant larvae or in Hsc70-4Δ19/Hsc70-4Δ11 or Hsc70-4Δ19/Hsc70-4Δ356 transheterozygotes are reduced to about half that of controls. EJP amplitudes are not reduced by a fatigue of evoked release, because responses sequentially recorded at 0.2 Hz do not progressively diminish. Evoked but not spontaneous synaptic transmission is thermo-intolerant in mutants. After increasing the temperature from 23oC to 30oC, homozygous mutants show a complete loss of EJP amplitudes after 20 minutes when stimulated at 0.2 Hz.After cooling to 23oC, at least a partial recovery of EJPs is always observed within 11 minutes. At a normal electrotonic stimulation intensity, application of Tetrodotoxin abolishes nerve-evoked EJPs in mutants as in wild-type. Subsequent stimulation with intensities 10 fold over threshold then elicit EJPs both in mutants and wild-type. However such electrotonically elicited EJPs are still reduced in mutants to about half that of controls, thus indicating a defect downstream of nerve excitation. Analysis of spontaneous neurotransmitter release reveals no postsynaptic defects at mutant neuromuscular junctions (NMJs). Mutant NMJs also have no abnormalities in their gross morphology. The quantal content of evoked release in homozygotes and Hsc70-4Δ19/Hsc70-4Δ11 or Hsc70-4Δ19/Hsc70-4Δ356 transheterozygotes is reduced to a third to a half of control. The estimated quantal content in homozygotes is significantly reduced for each examined Ca<up>2+</up>e compared to controls. Nerve evoked cytosolic Ca2+ levels during stimulation attain similar or higher values in homozygous mutants than in control boutons. The loss in neurotransmitter release in mutants is caused by a defect in exocytosis downstream of ca2+ entry. Synaptic vesicle recycling at mutant NMJs is normal. Ultrastructural studies of 3rd instar larval NMJs show that mutant terminals contain a large number of clear synaptic vesicles similar to controls. As in controls clathrin coated vesicles are not detectable. During repetitive stimulation, the evoked release are not depressed, suggesting that the number of releasable vesicles does not limit exocytosis in homozygous mutants. Increasing amounts of external and internal (residual) Ca2+ counteract a reduced probability of vesicle fusion in mutants caused by a decreased sensitivity of the Ca2+ signalling pathway triggering function.
Sgt56 ; Hsc70-4Δ19/Df(3R)BSC471 double mutant larvae show the same decreased level of endosomal microautophagy and significant decrease in mean excitatory junctional current amplitude at the neuromuscular junction as seen in Hsc70-4Δ19/Df(3R)BSC471 single mutants.
The increase in the number of multivesicular bodies (MVBs) and in the number of intraluminal vesicles per MVB area seen at the neuromuscular junction in Sgt56 larvae is suppressed in Sgt56 ; Hsc70-4Δ19/Df(3R)BSC471 double mutant larvae.
Hsc70-4Δ19/Df(3R)BSC471 is rescued by Hsc70-4wt.Tag:HA
Hsc70-4Δ19 is rescued by Hsc70-4+t14
Hsc70-4Δ19/Df(3R)BSC471 is partially rescued by Hsc70-4D10N.Tag:HA
Hsc70-4Δ19/Df(3R)BSC471 is not rescued by Hsc70-43KA.Tag:HA
Expression of either Hsc70-4wt.T:Ivir\HA1 or Hsc70-4D10N.T:Ivir\HA1 completely rescues the decrease in endosomal microautophagy which is seen at the neuromuscular junction in Hsc70-4Δ19/Df(3R)BSC471 larvae.
Expression of Hsc70-43KA.T:Ivir\HA1 fails to rescue the decrease in endosomal microautophagy which is seen at the neuromuscular junction in Hsc70-4Δ19/Df(3R)BSC471 larvae.
Expression of Hsc70-4wt.T:Ivir\HA1 completely rescues the decrease in mean excitatory junctional current amplitude which is seen at the neuromuscular junction in Hsc70-4Δ19/Df(3R)BSC471 larvae.
Expression of either Hsc70-43KA.T:Ivir\HA1 or Hsc70-4D10N.T:Ivir\HA1 does not rescue the decrease in mean excitatory junctional current amplitude which is seen at the neuromuscular junction in Hsc70-4Δ19/Df(3R)BSC471 larvae.