Cloning and Expression of Bovine Sodium/Glucose Cotransporter SGLT2

Abstract

The second member of the Na(+)/glucose cotransporter family, SGLT2, is a low-affinity active glucose transporter. In humans, it is predominantly located on the apical membrane of the S1 and S2 segments of renal proximal convoluted tubules, and, thus, may be mainly responsible for the reabsorption of D-glucose from the glomerular filtrate. By BLAST searching the GenBank database, we found expressed sequence tag sequences of SGLT2 in the cDNA library of bovine mammary tissues, indicating its expression in bovine mammary gland. To facilitate study of the mechanism of glucose reabsorption in bovine kidneys in maintenance of glucose homeostasis of lactating cows and the potential role of SGLT2 in the mammary gland, we cloned bovine SGLT2 and examined the distribution of its mRNA expression in bovine tissues. The full length mRNA of bSGLT2 is 2275 bp, and is predicted to encode a protein of 673 amino acids, with a molecular weight of approximately 73 kDa. The deduced amino acid sequence of bovine SGLT2 is 91, 90, 91, and 90% identical to human, rabbit, mouse, and rat SGLT2, respectively, and is 58 and 48% identical to bovine SGLT1 and SGLT5, respectively. The sequence of bSGLT2 contains several characteristically conserved sodium:solute symporter family signatures. Analysis of current bovine genomic data indicates that the bovine SGLT2 gene may consist of 14 exons. The major in vitro transcription and translation product of bovine SGLT2 cDNA migrated at an apparent molecular weight of 55 kDa. The SGLT2 mRNA was detected predominantly in bovine kidney as a 2.3-kb transcript, and at lower levels in all other bovine tissues examined, including the mammary gland, liver, lung, spleen, intestine, and skeletal muscle, as a 3.0-kb transcript. Expression of SGLT2 mRNA in bovine mammary gland increased more than 10-fold from late pregnancy to early lactation, similar to SGLT1. This indicates that SGLT2 may play a role in milk synthesis in the lactating mammary gland.

Functional Properties and Genomics of Glucose Transporters

“…- The members of the SGLT family also share considerable homology among the proteins (21-70% amino acid identity to SGLT1) and contain several conserved sodium:solute symporter family signatures that are characteristics of this family (GenBank accession numbers PS00456, PS00457 and PS50283). The SGLTs are predicted to have 14-15 transmembrane helices [3,16,17]. This second structural model is supported by experimental data of glycosylation studies, antibody tagging, mass spectrometry and other methods [3].…”

“…The hydrophilic N-terminus is located on the extracellular surface of the plasma membrane. There are several consensus sites for N-linked glycosylation [16,17], however, glycosylation appears not to be required for SGLT1 activity in contrast to GLUT1 [3].…”

Expression of Na+-d-glucose cotransporter SGLT2 in rodents is kidney-specific and exhibits sex and species differences

“…lated proteins, and thus indicates the possibility of misinterpretation of immunocytochemical staining. Extrarenal expression of Sglt2 mRNA in several species has been previously proposed in kidney, liver, lung, spleen, eye, and mammary gland of several species (12,25,41,51,54,57); however, the cellular location of SGLT2 protein has not been determined. A more recent study with quantitative PCR analysis indicated a highly kidney-specific expression of Sglt2 mRNA in humans (13).…”

“…In rat, high concentrations of Sglt2 mRNA were found in the renal proximal tubule S1 segments and less in the S2 segments, whereas the S3 segments were negative (54). In bovine, low concentrations of Sglt2 mRNA were also observed in small intestine, liver, lung, spleen, mammary gland, skeletal muscle, and ciliary epithelium (12,56,57). In humans, by real-time PCR the highest concentration of Sglt2 mRNA was observed in kidneys, but a limited expression was also detected in various extrarenal organs/tissues including small intestine, liver, brain, prostate, and testis (59), whereas recent studies indicated a highly kidney-specific expression of Sglt2 mRNA (13).…”

“…NaGLT1, a low-affinity Na ϩ -D-glucose cotransporter that does not trans-port D-galactose, has been cloned from rat and is highly expressed in kidney, where it is located in the BBM of proximal tubules (22). SGLT2 has been cloned from human (25,51), pig (26,31), rat (54), mouse (41), and bovine (57). The distribution of SGLT2 was mainly investigated on the mRNA level by Northern blot analysis and in situ hybridization.…”