The Rst-Neph Family of Cell Adhesion Molecules in Gallus gallus

Abstract

The Rst-Neph family comprises an evolutionarily conserved group of single-pass transmembrane glycoproteins that belong to the immunoglobulin superfamily and participate in a wide range of cell adhesion and recognition events in both vertebrates and invertebrates. In mammals and fish, three Rst-Neph members, named Neph1–3, are present. Besides being widely expressed in the embryo, particularly in the developing nervous system, they also contribute to the formation and integrity of the urine filtration apparatus in the slit diaphragm of kidney glomerular podocytes, where they form homodimers, as well as heterodimers with Nephrin, another immunoglobulin-like cell adhesion molecule. In mice, absence of Neph1 causes severe proteinuria, podocyte effacement and perinatal death, while in humans, a mutated form of Nephrin leads to congenital nephrotic syndrome of the Finnish type. Intriguingly, neither Nephrin nor Neph3 are present in birds, which nevertheless have typical vertebrate kidneys with mammalian-like slit diaphragms. These characteristics make, in principle, avian systems very helpful for understanding the evolution and functional significance of the complex interactions displayed by Rst-Neph proteins. To this end we have started a systematic study of chicken Neph embryonic and post-embryonic expression, both at mRNA and protein level. RT-qPCR mRNA quantification of the two Neph paralogues in adult tissues showed that both are expressed in heart, brain, and retina. Neph1 is additionally present in kidney, liver, pancreas, lungs, and testicles, while Neph2 mRNA is barely detected in kidney, testicles, pancreas and absent in liver and lungs. In embryos, mRNA from both genes can already be detected at as early as stage HH14, and remain expressed until at least HH28. Finally, we used a specific antibody to examine the spatial dynamics and subcellular distribution of ggNeph2 between stages HH20–28, particularly in the mesonephros, dermomyotomes, developing heart, and retina.

Keywords::

• chicken
• immunocytochemistry
• IRM proteins
• Neph1
• Neph2
• RT-qPCR

ACKNOWLEDGEMENTS

We would like to thank Dr. Roberta Ribeiro Costa Rosales and Elizabete Rosa Milani of Department of Cell and Molecular Biology Multiuser Laboratories of Confocal and Multiphoton Microscopy for help with the Confocal Microscope Leica SP5 and Vani Maria Alves of the Histology Facility of the same department for her invaluable help with the histological preparations. This work is supported by grants from FAPESP, CAPES, CNPq and FAEPA.

Declaration of interest: The authors report no declarations of interest. The authors alone are responsible for the content and writing of the paper.