Identification of three prominin homologs and characterization of their messenger RNA expression in Xenopus laevis tissues

Purpose Prominin is a family of pentaspan transmembrane glycoproteins. They are expressed in various types of cells, including many stem/progenitor cells. Prominin-1 plays an important role in generating and maintaining the structure of the photoreceptors. In this study, we identified three prominin homologs in Xenopus laevis, a model animal widely used in vision research, and characterized their messenger RNA (mRNA) expression in selected tissues of this frog. Methods Reverse-transcription PCR (RT–PCR) and rapid amplification of cDNA ends (RACE) were used to isolate cDNAs of prominin homologs. Semiquantitative RT–PCR was used to measure the relative expression levels of mRNAs of the three prominin homologs in four X. laevis tissues, specifically those of the retina, brain, testis, and kidney. Sequences of prominin homologs were analyzed with bioinformatic software. Results We isolated cDNAs of three prominin homologs from X. laevis tissues and compared their sequences with previously described prominin-1, 2, and 3 sequences from other species using phylogenetic analysis. Two of these homologs are likely to be the X. laevis orthologs of mammalian prominin-1 and 2, respectively, while the third homolog is likely to be the X. laevis ortholog of prominin-3, which was only found in nonmammalian vertebrates and the platypus. We identified alternatively spliced exons in mRNAs of all three prominin homologs. Similar to mammalian prominin-1, we found that exons 26b, 27, and 28a of the X. laevis prominin-1 gene are alternatively spliced, and that the splice isoforms of mRNA show tissue-specific expression profiles. We found that prominin-1 was the most abundant homolog expressed in the retina, brain, and testis, while prominin-3 was the most abundant homolog in the kidney. The expression level of prominin-2 was the lowest of the three prominin homologs in all four examined tissues of this frog. Conclusions Our results suggest that the mRNAs of prominin homologs are expressed in many tissues of X. laevis, but differ in their expression levels and mRNA splicing. Prominin-1 is the most abundant of the three prominin homologs expressed in the frog retina

Dysfunction of Heterotrimeric Kinesin-2 in Rod Photoreceptor Cells and the Role of Opsin Mislocalization in Rapid Cell Death

Loss of kinesin-2 function causes rapid death of rod photoreceptors. The cell death is dependent on the expression of opsin, which first accumulates along the route to the outer segment, but not on signaling by opsin-arrestin complexes or by light activation; the key element appears to be the accumulation of excessive protein in the wrong place

Immunocytochemistry at the Electron-Microscopic Level

Since the early slxtles, a number of reports have been published concerning immunocytochemical techniques for the detection of intracellular constituents at the electron microscope level. The· initial studies were made possible by the introduction by Singer of ferritin-tagged antibodies which could be visualized in the electron microscope. From all the procedures described later, immunocytochemical methods at the electron microscope can be classified into two main categories: 1) preembedding staining and 2) postembedding staining techniques

Prominin-1 Localizes to the Open Rims of Outer Segment Lamellae in Xenopus laevis Rod and Cone Photoreceptors

Antibodies specific for the N or C termini of xlProminin-1 labeled the open rims of lamellae of cone outer segments (COS) and the open lamellae at the base of rod outer segments (ROS). By contrast, anti-peripherin-2/rds antibody, Xper5A11, labeled the closed rims of cone lamellae adjacent to the ciliary axoneme and the rims of the closed ROS disks

On the absence of ubiquitous structural protein subunits in biological membranes

Although biological membranes exhibit a great diversity of functions, a number of general features appear to be similar in all membranes. In order to obtain a general concept of membrane structure and function, attempts have been made to find protein subunits responsible for the general structure of membranes, and numerous reports on such “structural proteins” appeared throughout the past decade. It was assumed that the structural proteins were necessary to maintain the membrane structure, whereas the special functions of each membrane were maintained by other proteins, e.g., enzymes

On the absence of ubiquitous structural protein subunits in biological membranes

Although biological membranes exhibit a great diversity of functions, a number of general features appear to be similar in all membranes. In order to obtain a general concept of membrane structure and function, attempts have been made to find protein subunits responsible for the general structure of membranes, and numerous reports on such “structural proteins” appeared throughout the past decade. It was assumed that the structural proteins were necessary to maintain the membrane structure, whereas the special functions of each membrane were maintained by other proteins, e.g., enzymes