Neuron-wide RNA transport combines with netrin-mediated local translation to spatially regulate the synaptic proteome
- University of California, Los Angeles, United States
Figures
Figure 1 with 1 supplement
rRNA targets equally well to SN branches contacting target and non-target MNs.
A bifurcated SN was cultured with an L7 target MN and an L11 non-target MN (A) for 3 days. Alexa647 (cyan) was microinjected into the SN before imaging to visualize SN processes. Cultures were …
Figure 1—figure supplement 1
28S rRNA targets equally well to SN branches contacting target and non-target MNs.
A bifurcated SN was cultured with an L7 target MN and an L11 non-target MN (A) for 3 days. Alexa647 (cyan) was microinjected into the SN before imaging to visualize SN processes. Cultures were …
Figure 2 with 2 supplements
Sensorin mRNA targets equally well to SN branches contacting target and non-target MNs.
A bifurcated SN was cultured with an L7 target MN and a L11 non-target MN (A) for 3 days. Alexa647 (cyan) was microinjected into the SN before imaging to visualize SN processes; in (A), Alexa 546 …
Figure 2—figure supplement 1
Sensorin mRNA distribution in DIV3 SN-MN cultures and localization in DIV1 SN-MN cultures.
The coefficient of variation for sensorin mRNA (A) indicates that the pattern of sensorin mRNA distribution is distinct in distal SN branches contacting L7 target MN, as previously described in Lyles…
Figure 2—figure supplement 2
β-Thymosin mRNA localizes equally well to SN branches contacting target and non-target MNs.
A bifurcated SN was cultured with an L7 target MN and an L11 non-target MN (A) for 3 days. Alexa647 (cyan) was microinjected into the SN before imaging to visualize SN processes. Cultures were …
Figure 3
EF1α mRNA is induced by 5x5HT and is delivered throughout the neuronal arbor.
A bifurcated SN was cultured with an L7 target MN and an L11 non-target MN (A and B) for 3 days. Alexa647 (cyan) was microinjected into the SN before imaging to visualize SN processes. On day 3, …
Figure 4 with 1 supplement
The RNA binding protein Aplysia Staufen targets equally well to SN neurites contacting L7 target and L11 non-target MNs.
(A) C-terminally tagged dendra2-tagged Aplysia Staufen (ApStaufen) was microinjected together with Alexafluor 647 (cyan) into a cultured bifurcated SN contacting an L7 target MN and an L11 …
Figure 4—figure supplement 1
The RNA binding protein Aplysia Staufen targets equally well to stimulated- and unstimulated-SN processes after local perfusion.
(A) and (B) bifurcated sensory neurons (SNs) were cultured with two target LFS motor neurons (MNs). C-terminally tagged dendra2-tagged Aplysia Staufen (ApStaufen) was expressed in the SN, which was …
Figure 5 with 1 supplement
Local translation is greater in SN neurites contacting L7 target MNs than in neurites contacting L11 non-target MNs.
(A) A bifurcated SN was cultured with an L7 target MN and an L11 non-target MN for 3 days. Alexa647 was microinjected into the SN and served as a volume filling control for analysis of …
Figure 5—figure supplement 1
Local translation is enriched in SN branches contacting L7 MN.
A bifurcated SN was cultured with an L7 target MN and an L11 non-target MN (photomicrographs in A, D, and G) for 3 days. Alexa dye 647 (cyan) was microinjected into the SN before imaging (A, D, and G…
Figure 6
Global translation is also enriched in SN branches contacting L7 MN.
A bifurcated SN was cultured with an L7 target MN and an L11 non-target MN (A) for 3 days. Alexa488 (green) was microinjected into the SN as a volume control for imaging (A). Cultured neurons were …
Figure 7 with 1 supplement
Netrin-1 increases protein synthesis in SNs and strength of SN-target MN synapses.
(A) A bifurcated sensory neuron was cultured with an L7 target MN and an L11 non-target MN for 3 days. Alexa647 was microinjected into SNs and served as a volume filling control for the analysis of …
Figure 7—figure supplement 1
Netrin-1 increases local translation in soma-free SN processes in SN-MN cultures.
SN-MNs were cultured for 3 days and the SN soma (denoted by dotted circle in A) was mechanically removed with sharp glass electrode. 12–18 hr later, cultures were incubated with recombinant Fc human …
Figure 8
Netrin-1 binds DCC to promote SN translation and to increase strength of SN-MN synapses.
(A) and (B): immunocytochemistry (ICC) of isolated SNs (3DIV) reveals an increase in sensorin immunoreactivity with recombinant Fc human netrin-1 incubation (250 ng/ml for 24 hr), but no change from …
Figure 9
Overexpression of Aplysia netrin-1 increases translation and synaptic strength in SN-MN synapses.
SN-LFS MNs were cultured for 3 days (A–C). The SN was microinjected with Alexa647 (blue) and the MN with a vector expressing Aplysia netrin-1 (ApNetrin-1) c-terminally tagged with dendra2 (green, A).…
Figure 10 with 1 supplement
ApNetrin-1 undergoes calcium-dependent release from MN and binds SN.
SN-LFS MNs were cultured for 2 days, and the SN was microinjected with a vector expressing ApDCC c-terminally tagged with mCherry (red) and the MN with Alexa647 (blue) and with a vector expressing …
Figure 10—figure supplement 1
Incubation with anti-DCC antibodies inhibits binding of MN-expressed ApNetrin-1 to SNs.
SN-LFS MNs were cultured for 2 days, and the SN was microinjected with a vector expressing ApDCC c-terminally tagged with mCherry (red) and the MN with Alexa647 (blue) and with a vector expressing …
