Submitted on September 25, 2007
Accepted on November 28, 2007
Zebrafish Ae2.2 encodes a second Slc4a2 anion exchanger
Boris E. Shmukler1, Jeffrey S. Clark2, Ann Hsu3, David H. Vandorpe1, Andrew K. Stewart1, Christine E Kurschat4, Seong-Kyu Choe5, Yi Zhou6, Julio Amigo5, Barry H. Paw5, and Seth L. Alper1*
1 Molecular and Vascular Medicine and Renal Unit, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States; Medicine, Harvard Medical School, Boston, Massachusetts, United States
2 Medicine, Harvard Medical School, Boston, Massachusetts, United States; Molecular and Vascular Medicine and Renal Unit, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
3 Molecular and Vascular Medicine and Renal Unit, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States; Neuroscience, Mt. Holyoke College, S. Hadley, Massachusetts, United States
4 Molecular and Vascular Medicine and Renal Unit, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
5 Hematology Division, Brigham and Women's Hospital, Boston, Massachusetts, United States; Medicine, Harvard Medical School, Boston, Massachusetts, United States
6 Hematology Division, Children's Hospital Boston, Boston, Massachusetts, United States; Pediatrics, Harvard Medical School, Boston, Massachusetts, United States
* To whom correspondence should be addressed. E-mail: salper{at}bidmc.harvard.edu.
The genome of zebrafish D. rerio encodes two unlinked genes equally closely related to the SLC4A2/AE2 anion exchanger genes of mammals. One of these is the recently reported zebrafish Ae2 gene (Shmukler et al., Am J Physiol Renal Physiol 289:R835, 2005), now called Ae2.1. We now report the structural and functional characterization of the second zebrafish AE2 gene product, Ae2.2. The Ae2.2 gene of zebrafish LG24 encodes a polypeptide of 1232 aa in length, sharing 70% amino acid identity with zebrafish Ae2.1 and 67% identity with mouse AE2a. Zebrafish Ae2.2 expressed in Xenopus oocytes encodes a 135 kDa polypeptide that mediates bidirectional, DIDS-sensitive Cl-/Cl- exchange and Cl-/HCO3- exchange. Ae2.2-mediated Cl-/Cl- exchange is cation-independent, voltage-insensitive, and electroneutral. Acute regulation of anion exchange mediated by Ae2.2 includes activation by NH4+ and independent inhibition by acidic intracellular pH (pHi) and by acidic extracellular pH (pHo). In situ hybridization reveals low-level expression of Ae2.2 mRNA in zebrafish embryo, most notably in posterior tectum, eye, pharynx, epidermal cells, and axial vascular structures, without notable expression in the Ae2.1-expressing pronephric duct. Knockdown of Ae2.2 mRNA, of Ae2.1 mRNA, or of both with nontoxic levels of N-morpholino-oligomers produced no grossly detectable morphological phenotype, and preserved normal structure of the head and the pronephric duct at 24 hpf.
