Identification and expression of XRTN1-A and XRTN1-C in Xenopus laevis

Abstract

Reticulon1, also known as neuroendocrine-specific protein, belongs to the reticulon (RTN) family, whose members possess a conserved reticulon domain and are associated with the endoplasmic reticulum (ER) membrane. Here, we report cloning and expression of Xenopus homologues of Reticulon1-A (XRTN1-A) and -C (XRTN1-C). XRTN1-A and -C contain an open reading frame of 752 and 207 amino acids, respectively, each containing a conserved reticulon domain. Sequence analysis shows that XRTN1 proteins have an ER membrane retention signal and four putative membrane-spanning domains. Reverse transcription-polymerase chain reaction and whole-mount in situ hybridization showed that XRTN1-A is expressed in early neural precursors and differentiating neuronal populations, including the trigeminal placode, olfactory placode, lateral line placode, and otic vesicle. XRTN1-C is expressed in the developing brain and spinal cord. We found that XRTN1-C protein is localized to the ER of Xenopus and mammalian cells and the granules in neurites of primary neurons of the Xenopus spinal cord and rat hippocampus. We also showed that XRTN1-C protein is detected in the heavy membrane fraction, which contains lysosomal and ER-resident proteins, as well as in the nucleus and polysomal fractions of the Xenopus embryo. Finally, we showed that thyroid hormone specifically down-regulates XRTN1-A mRNA in the head of premetamorphic Xenopus tadpoles. Our work characterizes the intracellular roles of XRTN1 during Xenopus neural development.