Organ morphogenesis requires cellular shape changes and tissue rearrangements that occur in a precisely timed manner. Here, we show that zebrafish heart and soul (Has)/protein kinase C iota (PRKCi) is required tissue-autonomously within the myocardium for normal heart morphogenesis and that this function depends on its catalytic activity. In addition, we demonstrate that nagie oko (Nok) is the functional homolog of mammalian protein associated with Lin-seven 1 (Pals1)/MAGUK p55 subfamily member 5 (Mpp5), and we dissect its earlier and later functions during myocardial morphogenesis. Has/PRKCi and Nok/Mpp5 are required early for the polarized epithelial organization and coherence of myocardial cells during heart cone formation. Zygotic nok/mpp5 mutants have later myocardial defects, including an incomplete heart tube elongation corresponding with a failure of myocardial cells to correctly expand in size. Furthermore, we show that nok/mpp5 acts within myocardial cells during heart tube elongation.Together, these results demonstrate that cardiac morphogenesis depends on the polarized organization and coherence of the myocardium, and that the expansion of myocardial cell size contributes to the transformation of the heart cone into an elongated tube.KEY WORDS: Organ morphogenesis, prkci, Myocardium, heart and soul, nagie oko, mpp5, pals1, Zebrafish Development 133,[107][108][109][110][111][112][113][114][115]

Abstract-Many vertebrate organs are derived from monolayered epithelia that undergo morphogenesis to acquire their shape. Whereas asymmetric left/right gene expression within the zebrafish heart field has been well documented, little is known about the tissue movements and cellular changes underlying early cardiac morphogenesis. Here, we demonstrate that asymmetric involution of the myocardium of the right-posterior heart field generates the ventral floor, whereas the noninvoluting left heart field gives rise to the dorsal roof of the primary heart tube. During heart tube formation, asymmetric left/right gene expression within the myocardium correlates with asymmetric tissue morphogenesis. Disruption of left/right gene expression causes randomized myocardial tissue involution. Time-lapse analysis combined with genetic analyses reveals that motility of the myocardial epithelium is a tissue migration process. Our results demonstrate that asymmetric morphogenetic movements of the 2 bilateral myocardial cell populations generate different dorsoventral regions of the zebrafish heart tube. Failure to generate a heart tube does not affect the acquisition of atrial versus ventricular cardiac cell shapes. Therefore, establishment of basic cardiac cell shapes precedes cardiac function. Together, these results provide the framework for the integration of single cell behaviors during the formation of the vertebrate primary heart tube. (Circ Res. 2008;102:e12-e19.)Key Words: heart tube Ⅲ cell polarity Ⅲ protein kinase C iota Ⅲ left-right asymmetry Ⅲ southpaw Ⅲ nagie oko H eart development in vertebrates involves the fusion of 2 myocardial progenitor fields at the embryonic midline. These heart fields derive from the left and right lateral plate mesoderm (LPM). 1 In zebrafish, the fusion of the 2 heart fields forms the heart cone, a central flat disc that is subsequently transformed into the primary heart tube that generates a 2-chambered heart with an anterior atrium and a posterior ventricle, which initiates circulation at 24 hours post fertilization (hpf). [2][3][4] Morphogenetic processes and tissue dynamics required for heart cone-to-tube transition are not well understood. Previous studies have described the asymmetric leftward movement of the primary heart tube (after 24 hpf), followed by the looping of the heart at 36 hpf, processes that depend on the left/right (L/R) signaling pathway and transform the linear heart tube into a looped heart with distinct bean-shaped heart chambers. 5 A key player in the hierarchy of the L/R signaling pathway is the nodal-related gene southpaw (spaw), which also affects the correct expression of downstream genes including pitx2, lefty1, and lefty2. 6 Combinatorial gene expression patterns of L/R signaling genes have been described within the heart cone. 7 However, whether this L/R asymmetric gene expression is underlying asymmetric cell behaviors is currently unknown.Myocardial precursor cells acquire a polarized epithelial morphology, which is a prerequisite for normal heart developme...

Megalin/LRP2 is an endocytic receptor in the proximal tubules of the mammalian kidney that plays a central role in the clearance of metabolites from the glomerular filtrate. To establish a genetic model system for elucidation of molecular components of this retrieval pathway, we characterized orthologous transport processes in the zebrafish. We show that expression of megalin/LRP2 and its co-receptor cubilin is conserved in the larval zebrafish pronephros and demarcates a segment of the pronephric duct that is active in clearance of tracer from the ultrafiltrate. Knock-down of megalin/LRP2 causes lack of Rab4-positive endosomes in the proximal pronephric duct epithelium and abrogates apical endocytosis. Similarly, knock-down of the megalin/LRP2 adaptor Disabled 2 also blocks renal clearance processes. These results demonstrate the conservation of the megalin/LRP2 retrieval pathway between the larval zebrafish pronephros and the mammalian kidney and set the stage for dissection of the renal endocytic machinery in a simple model organism. Using this model system, we provide first genetic evidence that renal tubular endocytosis and formation of endosomes is a ligand-induced process that crucially depends on megalin/LRP2 activity.

Retinal lamination is known to depend on cell polarity and localized signaling. In vertebrates, atypical Protein Kinase C proteins, aPKCλ/ι and aPKCζ, are essential for apical-basal cell polarity. However, it is not known to what extent functional redundancy has precluded a comprehensive functional characterization of aPKC signaling during vertebrate retinogenesis. Here, we show that aPKCs λ and ζ are functionally redundant for multiple aspects of retinogenesis including mitotic division location and orientation, cell-type positioning, and retinal pigment epithelial (RPE) and photoreceptor cell morphogenesis. Genetic mosaic analyses demonstrate a cell-autonomous requirement of aPKCs for RPE and photoreceptor development, and a cell-non-autonomous function that is intrinsic to the neural retina for cell-type positioning. Our observations uncover a previously unappreciated involvement of aPKCζ during zebrafish retinogenesis and suggest that aPKC signaling primes the retinal environment for appropriate cell migration of post-mitotic progenitor cells, but is not essential for correct cell-type specification.