title: Expression and localization of CERKL in the mammalian retina, its response to light-stress, and relationship with NeuroD1 gene.
abstract: Mutations in the Ceramide kinase like (CERKL) gene are associated with retinitis pigmentosa (RP26) and cone-rod dystrophy. CERKL is homologous to Ceramide kinase (CERK), and its function is still unknown. The purpose of this study was to test the expression and distribution of this gene and its protein in rat and in mouse tissues, in light-stressed rat retinas and in the retinas of NeuroD1 knock-out mice to understand the role of CERKL in the retina. Expression of Cerkl and Cerk mRNA was determined by quantitative RT-PCR. Expression of the protein was determined by Western blotting with anti-CERKL antibody. Localization of the protein was determined by using immunofluorescence microscopy. With qRT-PCR, we revealed that the relative mRNA expression of Cerkl was the highest in the retina among all the rat tissue tested; it was >10-fold higher than in the brain. On the other hand, Cerk has ubiquitous expression and its relative abundance is >2 fold of Cerkl in the retina. Cerkl was expressed minimally in the developing mouse eyes and reached a peak at retinal maturity at 2 months. Western blots of retinal tissues revealed two major CERKL protein bands: 59 kDa (C1) and 37 kDa (C2). However, only C2 CERKL was found in the rat retinal rod outer segment (ROS) at level of that was not changed in light vs. dark adaptation. In the light-stressed retina, expression of Cerkl mRNA increased significantly, which was reflected in only on C2 CERKL protein. The CERKL protein localized prominently to the ganglion cells, inner nuclear layers (INL), retinal pigment epithelial (RPE) cells, and photoreceptor inner segments in the retinal sections. Nuclear localization of CERKL was not affected in RPE, INL and the ganglion cell layers in the light-stressed retina; however, the perinuclear and outer segment locations appear to be altered. In the NeuroD1 knock-out mouse retina, the expression of Cerkl mRNA and protein decreased and that decrease also pertains to C2 CERKL. In conclusion, the retina had the highest level of Cerkl mRNA and protein expression, which reached its maximum in the adult retina; CERKL localized to ROS and RPE cells and the light-adaptation did not change the level of CERKL in ROS; light-stress induced Cerkl expression in the retina; and its expression decreased in NeuroD1 knock-out retina. Thus, CERKL may be important for the stress responses and protection of photoreceptor cells.
authors.affiliation: Department of Ophthalmology, OUHSC, Dean A. McGee Eye Institute, Oklahoma City, OK 73104, United States. mmandal@ouhsc.edu
authors.forename: Nawajes A
authors.initials: NA
authors.lastname: Mandal
authors.forename: Julie-Thu A
authors.initials: JT
authors.lastname: Tran
authors.forename: Anisse
authors.initials: A
authors.lastname: Saadi
authors.forename: Abul K
authors.initials: AK
authors.lastname: Rahman
authors.forename: Tuan-Phat
authors.initials: TP
authors.lastname: Huynh
authors.forename: William H
authors.initials: WH
authors.lastname: Klein
authors.forename: Jang-Hyeon
authors.initials: JH
authors.lastname: Cho
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics*; D015153:Blotting, Western; D005786:Gene Expression Regulation / Q000502:physiology*; D008027:Light / Q000009:adverse effects*; D051379:Mice; D008807:Mice, Inbred BALB C; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011833:Radiation Injuries, Experimental / Q000209:etiology / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D060888:Real-Time Polymerase Chain Reaction; D012160:Retina / Q000196:embryology / Q000378:metabolism / Q000528:radiation effects*; D012162:Retinal Degeneration / Q000209:etiology / Q000235:genetics* / Q000378:metabolism; D055213:Retinal Pigment Epithelium / Q000378:metabolism; D012374:Rod Cell Outer Segment / Q000378:metabolism
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D017853:Phosphotransferases (Alcohol Group Acceptor); D012333:RNA, Messenger; C501008:Neurod1 protein, mouse; C530392:CerkL protein, mouse