Gattegno, Tamar et al. (2015) International Worm Meeting "AFF-1 fuses terminal branches non cell-autonomously during dendritic regeneration."

Oren-Suissa, Meital, Podbilewicz, Benjamin, Kravtsov, Veronika, Gattegno, Tamar

[International Worm Meeting, 2015]

We wish to reveal the basic requirement for dendritic regeneration after injury in order to better understand neuronal plasticity and maintenance. Our model is the PVDs, a pair of highly arborized mechanosensory neurons that are responsible for sensation of strong mechanical stimuli, temperature and posture. PVDs develop through a dynamic process forming its structural units termed "menorahs" that define the resolution of its receptive field (Oren-Suissa et al., 2010). Two cell-cell fusion proteins (EFF-1 and AFF-1) have been identified in C. elegans and they act to mediate different cell fusion events in the hypodermis, pharynx, vulva, uterus, and other organs. EFF-1 has a function to regenerate broken axons by auto-fusion (Ghosh-Roy et al., 2010) and to sculpt PVD dendrites by branch fusion and retraction during development (Oren-Suissa et al., 2010). AFF-1 mediates fusion events of cells whose fusion does not require EFF-1 e.g. anchor cell to utse syncytium and seam cells fusions (Sapir et al., 2007).To understand how dendrites regenerate following injury, we used a femtosecond laser to sever PVD dendrites and followed their regeneration in wild-type and mutant animals. We demonstrate that the reconnection of broken branches occurs through fusion using Kaede photoactivation. PVD dendritic regeneration process involves branch sprouting along with loss of self-avoidance of the menorahs thus allowing fusion of terminal branches to bypass the site of injury. As opposed to wild type, where most of PVD regeneration occurred through terminal branches fusion, in aff-1 mutants PVD regeneration occurred mainly through primary branch fusion. Our results suggest that AFF-1 fuses terminal branches to enable efficient regeneration.We tested whether AFF-1 acts cell autonomously to fuse broken dendrites using tissue-specific rescue experiments. Although AFF-1 expresses in sheath cells of chemosensory neurons in the head we did not observe its expression in the PVD, before or after injury. Moreover, following PVD regeneration of transgenic aff-1 null mutants that express AFF-1 only in the PVD compared to their siblings, we confirmed that AFF-1 can fuse terminal branches; however AFF-1 expression in the PVD could not elevate the number of fused menorahs to wild type level, thus implying that AFF-1 acts non cell-autonomously to reconnect terminal dendrites. This is in contrast to EFF-1, that is expressed in the PVD and acts cell-autonomously to sculpt PVD dendrites. We are currently analyzing PVD regeneration in aff-1 mutants that express AFF -1 in the seam cells and in the hypodermis.

Brar, Harjot Kaur et al. (2021) International Worm Meeting "Dendrite regeneration in PVD neuron is controlled by the RAC GTPase CED-10 and the RhoGEF TIAM-1"

Ghosh-Roy, Anindya, Dey, Swagata, Brar, Harjot Kaur, Pandey, Devashish, Bhardwaj, Smriti, Dey, Shirshendu

[International Worm Meeting, 2021]

Neurons are vulnerable to physical insults which compromise the integrity of both dendrites and axons. Unlike well characterized axon regeneration, our knowledge in dendrite regeneration is limited. To understand the mechanisms of dendrite regeneration, we used PVD neurons with stereotyped branched dendrites. Using femtosecond laser, we severed the primary dendrites and axon of this neuron. After the primary dendrite was severed near the cell body, we observed a sprouting of new branches from the cut site within 3 hours. By 24 hours, the primary dendrite regrew to cover the original territory in complex pattern unlike the uninjured dendrites. We quantified the regeneration in broadly two aspects- the territory covered and fusion phenomena (Oren-Suissa et al.,2017, Kravtsov et al.,2017). Axon injury causes a retraction of the severed end followed by a Dual leucine zipper kinase-1(DLK-1) dependent regrowth from the severed end or conversion of neighboring dendrite to axon. However, Dendrite regeneration was independent of DLK-1 and other conventional axon regeneration pathways including cAMP elevation, let-7 miRNA, Akt-1 and Phosphatidyl serine exposure/PS. Among various candidates tested, ced-10 mutants showed a defect in dendrite regeneration. It is a RAC GTPase involved in regulation of neuronal cytoskeleton and cell engulfment. Cell-specific rescue experiments suggest that cell-autonomous and epidermal expression of CED-10 is required for dendrite regrowth and fusion, respectively. Moreover, the PVD-specific expression of an activated version of CED-10 led to both increased branching and fusion. We ventured further to find the upstream players which can control the function of RAC GTPase in dendrite regeneration. We found out that the TIAM-1 RhoGEF is required for dendrite regeneration and the activated CED-10 can bypass the requirement of TIAM-1 in dendrite regrowth. Our work provides a framework for understanding the cellular mechanism of dendrite regeneration using PVD model. Reference: Oren-Suissa, M., T. Gattegno, V. Kravtsov and B. Podbilewicz, 2017 Extrinsic Repair of Injured Dendrites as a Paradigm for Regeneration by Fusion in Caenorhabditis elegans. Genetics 206: 215-230. Kravtsov, V., M. Oren-Suissa and B. Podbilewicz, 2017 The fusogen AFF-1 can rejuvenate the regenerative potential of adult dendritic trees by self-fusion. Development 144: 2364-2374.