Proteomic analysis of avian and mammalian intestinal brush border membrane

CAVIEDES-VIDAL, ENRIQUE; ANTONIO BRUN; MAGALLANES, MELISA E.; BARRETT-WILT, GREGORY A.; KARASOV WH

San Diego

Congreso; Comparative physiology: From Organisms to Omics in an Uncertain World; 2022

American Physiological Society

Guided by functional understanding, we tested several hypothesesregarding the abundance of key proteins of the intestinal brushborder membrane (BBM) of 4 avian species and 2 rodents: strictlygranivorous zebra finch (Taeniopygia guttata),granivorous/omnivorous domestic chickens (Gallus gallus) and housesparrows (Passer domesticus), insectivorous European starlings(Sturnus vulgaris), and omnivorous lab mice (Mus musculus) andrats (Rattus norvegicus). The whole intestine of 4 individuals ofeach species was harvested, enterocytes isolated, and BBM isolatesprepared. Proteins were solubilized, digested, andchromatographically separated. Peptides were analyzed by nano LCMS/MS. Raw MS/MS data were converted and used to search againstRefSeq amino acid sequence databases. We focus was on proteinsthat belong to the BBM of the enterocytes or tightly interact withthem. Thus, proteins were filtered applying the following GO terms:GO:0005902 - Microvillus, GO:0031526 - Brush border membrane,GO:0005903 - Brush border, GO:0016324 - Apical plasmamembrane and GO:1904659 - Glucose transmembrane transport.Although we expected that structural scaffold proteins (e.g ofcytoskeleton and apical cell junctions), which made up nearly half ofthe BBM proteins, would be evolutionarily conserved among all thespecies, we found marked proportional differences between birdsand mammals in abundances, defined as their respective spectranormalized by total spectra of all BBM protein detected. Furthermoremore, one of the most abundant structural proteins in the rodents,myosin heavy chain 14 (MHY14), was absent in the birds. This resultwas confirmed in a follow-up genomic analysis. As predicted forhydrolases, TREH was absent in birds, and abundances of others,e.g., alpha-glucosidases (AGs, SI, maltase) and peptidase (APN),correlated with their respective hydrolytic activity per unit intestine(measured previously). Also as predicted, the lowest ratio of AG:APNwas in the insectivorous starling, which has the lowest ratio ofcarbohydrate:protein in its diet. As predicted for transporters, theabundances of the active glucose transporter SLC5A1 (SGLT1) waslower in the perching birds than in rodents because they generallyrely more on paracellular sugar absorption than on transmembranetransporter mediated absorption. Interestingly, the abundances ofxenobiotic transporters were higher in birds than in rodents, butseveral competing hypotheses might predict that. Supported byU.S.-N.S.F (to WHK) and CONICET (to E.C.-V).