Although multiple determinants for establishing polarity in membranes of epithelial cells have been identified, the mechanism for maintaining apicobasal polarity is not fully understood. Here, we show that the conserved Hippo kinase pathway plays a role in the maintenance of apicobasal polarity in the developing intestine of Caenorhabditis elegans. We screened suppressors of the mutation in wts-1-the gene that encodes the LATS kinase homolog, deficiency of which leads to disturbance of the apicobasal polarity of the intestinal cells and to eventual death of the organism. We identified several alleles of yap-1 and egl-44 that suppress the effects of this mutation. yap-1 encodes a homolog of YAP/Yki, and egl-44 encodes a homolog of TEAD/Sd. WTS-1 bound directly to YAP-1 and inhibited its nuclear accumulation in intestinal cells. We also found that NFM-1, which is a homolog of NF2/Merlin, functioned in the same genetic pathway as WTS-1 to regulate YAP-1 to maintain cellular polarity. Transcriptome analysis identified several target candidates of the YAP-1-EGL-44 complex including TAT-2, which encodes a putative P-type ATPase. In summary, we have delineated the conserved Hippo pathway in C. elegans consisting of NFM-1-WTS-1-YAP-1-EGL-44 and proved that the proper regulation of YAP-1 by upstream NFM-1 and WTS-1 is essential for maintenance of apicobasal membrane identities of the growing intestine.

Edited by Tamas DalmayKeywords: MicroRNA Degradome sequencing MicroRNA-directed cleavage C. elegans a b s t r a c t Caenorhabditis elegans microRNAs (miRNAs) bind to partially complementary sequences in the 3 0 untranslated region of target mRNAs, resulting in translational repression through mRNA destabilization. High-throughput sequencing of RNA cleavage fragments was performed to directly detect miRNA-directed cleavage targets in adult stage C. elegans. From this analysis, we found that miR-249 directed the cleavage of the ZK637.6 transcript with extensive and evolutionarily conserved complementarity in nematode. In addition, expression of the ZK637.6 transcript was strongly dependent on the expression of miR-249. These findings may lead to a better understanding of miR-NA-mediated gene regulation in nematodes.

A fundamental question in neurodevelopmental biology is how flexibly the nervous system can change during development. To address this question of developmental plasticity, we analyzed the connectome of dauer, an alternative developmental stage of nematodes with physiological and behavioral characteristics remarkably distinct from other developmental stages. We reconstructed the complete chemical connectome of a dauer by manual volumetric reconstruction and automated synapse detection using deep learning. While the basic architecture of the nervous system was preserved, there were also structural changes in neurons, large or small, that were closely associated with changes in the connectivity, some of which in turn evoked dauer-specific behaviors such as nictation. Combining the connectome data and optogenetic experiments were enough to reveal dauer-specific neural connections for the dauer-specific behavior. Graph theoretical analyses showed higher clustering of motor neurons and more feedback connections from motor to sensory neurons in the dauer connectome, suggesting that the dauer connectome allows a quick response to an ever-changing environment. We suggest that the nervous system in the nematode, which can be extended to animals in general, has evolved to obtain the ability to respond to harsh environments by reversibly developing a connectome quantitatively and qualitatively differentiated from other developmental stages.

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