Our optimal visual performance starts with the correct parallel visual information processing in the retina which depends on precise retinal neural circuit formation. To achieve this, retinal neurons must accurately set up functional synapses which require precise alignment between the pre-synaptic neurotransmitter releasing site and the post-synaptic receptor. Increasing evidence suggests that cell adhesion molecules (CAMs)-mediated trans-synaptic complexes are critical for synapse formation and function, however, the mechanism how pre- synaptic releasing machinery coordinates with the post-synaptic receptor signaling complex at retinal ribbon synapses is largely unknown. We recently reported that the extracellular calcium channel auxiliary subunit ?2?4 is necessary for rod photoreceptor axonal elaboration and synapse formation and demonstrated that ?2?4 does so through controlling the synaptic targeting of ELFN1, a leucine-rich repeat (LRR) protein specifically expressed at rod synapse and interacts trans-synaptically with the post-synaptic mGluR6 receptor. We also showed that ?2?4 interacts with ELFN1 through the LRR domain, a conserved domain shared across many LRR proteins. Interestingly, we and other lab identified another LRR protein termed LRIT1, which specifically affects cone synaptic function. The objective of the proposed work is to determine whether it is a general mechanism that pre-synaptic calcium channel complex utilizes ?2?4 to coordinate with post-synaptic receptor through the facilitation of different LRR proteins.
In Aim 1, we will determine the functional role of ?2?4-ELFN1 interaction in rod synapse formation using in vivo electroporation combined with deletion mutagenesis.
In Aim 2, we will study how LRIT1 regulates cone synaptic function by testing whether LRIT1 affects pre-synaptic Cav1.4 channel activity through ?2?4 using electrophysiological recording.
In Aim 3, I will investigate whether synapses in the inner plexiform layer (IPL) also adopt similar trans-synaptic mechanism by studying how ?2?4 and LRIT1 affect RBC-AII amacrine synapse formation and function using the technique gained during the mentored phase. This proposal is innovative because no mechanistic studies has been done on ?2?4 protein despite its clearly important role in photoreceptor synapse formation and function. The proposed work is significant since results from this study will enable better understanding of how retinal neurons establish and maintain synaptic contact.
Malfunction of ?2?4 has been shown to cause retinal diseases such as night blindness and cone dystrophy. This project is aimed to elucidate detailed molecular mechanism underlying ?2?4?s function in retina and to advance our understanding on the pathology of night blindness and cone dystrophy for better therapeutics development. The results from the proposed research will also provide valuable molecular insights into retinal synapse formation and function and hence facilitate the optimization of the current transplantation therapy of which the biggest challenge is to re-establish the connectivity between the transplanted photoreceptors and the host retinal neurons. !
