Intestinal ribosomal p70(S6K) signaling is increased in piglet rotavirus enteritis.
Recent identification of the mammalian target of rapamycin (mTOR) pathway as an amino acid-sensing mechanism that regulates protein synthesis led us to investigate its role in rotavirus diarrhea. We hypothesized that malnutrition would reduce the jejunal protein synthetic rate and mTOR signaling via its target, ribosomal p70 S6 kinase (p70(S6K)). Newborn piglets were artificially fed from birth and infected with porcine rotavirus on day 5 of life. Study groups included infected (fully fed and 50% protein calorie malnourished) and noninfected fully fed controls. Initially, in "worst-case scenario studies," malnourished infected piglets were killed on days 1, 3, 5, and 11 postinoculation, and jejunal samples were compared with controls to determine the time course of injury and p70(S6K) activation. Using a 2 x 2 factorial design, we subsequently determined if infection and/or malnutrition affected mTOR activation on day 3. Western blot analysis and immunohistochemistry were used to measure total and phosphorylated p70(S6K); [(3)H]phenylalanine incorporation was used to measure protein synthesis; and lactase specific activity and villus-crypt dimensions were used to quantify injury. At the peak of diarrhea, the in vitro jejunal protein synthetic rate increased twofold (compared with the rate in the uninfected pig jejunum), concomitant with increased jejunal p70(S6K) phosphorylation (4-fold) and an increased p70(S6K) level (3-fold, P < 0.05). Malnutrition did not alter the magnitude of p70(S6K) activation. Immunolocalization revealed that infection produced a major induction of cytoplasmic p70(S6K) and nuclear phospho-p70(S6K), mainly in the crypt. A downregulation of semitendinosus muscle p70(S6K) phosphorylation was seen at days 1-3 postinoculation. In conclusion, intestinal activation of p70(S6K) was not inhibited by malnutrition but was strongly activated during an active state of mucosal regeneration.
Rhoads
J Marc
JM
Department of Pediatrics, Ochsner Clinic Foundation and Ochsner Children's Research Institute, New Orleans, Louisiana, USA. j.marc.rhoads@uth.tmc.edu
Corl
Benjamin A
BA
Harrell
Robert
R
Niu
Xiaomei
X
Gatlin
Lori
L
Phillips
Oulayvanh
O
Blikslager
Anthony
A
Moeser
Adam
A
Wu
Guoyao
G
Odle
Jack
J
D000596:Amino Acids / Q000097:blood; D000818:Animals; D000831:Animals, Newborn; D000956:Antigens, Viral / Q000032:analysis; D001835:Body Weight; D003967:Diarrhea / Q000378:metabolism / Q000473:pathology; D002149:Energy Intake; D004751:Enteritis / Q000097:blood / Q000378:metabolism* / Q000821:virology; D020895:Enterocytes / Q000737:chemistry / Q000378:metabolism; D005243:Feces / Q000737:chemistry; D007413:Intestinal Mucosa / Q000737:chemistry / Q000378:metabolism; D007583:Jejunum / Q000737:chemistry / Q000378:metabolism*; D043322:Lactase / Q000378:metabolism; D044342:Malnutrition / Q000378:metabolism / Q000503:physiopathology; D018482:Muscle, Skeletal / Q000378:metabolism; D009130:Muscle, Smooth / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000502:physiology; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000032:analysis / Q000378:metabolism*; D012400:Rotavirus Infections / Q000097:blood / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D034421:Sus scrofa; D058570:TOR Serine-Threonine Kinases
D000596:Amino Acids; D000956:Antigens, Viral; D011494:Protein Kinases; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; D043322:Lactase