Sites of protein synthesis as shown by radioautography after administration of S³⁵–labelled methionine

Abstract

and Oniocrdlg o f Waxkington, Seallle, Wmhinglon THIRTY-TTIBEE FIQURERSITES OF PUOTEII SYNTHESIS 227 yeast was added t,o 9 parts of the prepared diet formula arid then the expected weight gain was observed ( fig. 1). A few of the animals w-cre used in pilot expcrimcnts. The others werc treated as indicated below.'For radioa2~ictogru,phic observu.tions, DL-methionine-S35 (specific activity, 1 mc per 10.3 mg) 1va.s injoctcxi subcutaneously, 7.7 vc (0.0793 mg of methionine) per gram of body weight, 4 weeks after surgery. Injecti.ons were administered to three coat.rol. ruts which were sacrificed 4, 24 and 36 hours later respectively, and to t.wo rats from each of tlie tliyroidcct.omierd and thyroxine-tren,ted-thyroidectomized groups, which were sacrificed 4 and 36 hours latcr r.cspcc.t.ively. The tinimals were mc.st.l.ietizei1 with ether and killed by withdrawing a1.l the !)lootl oMainable from thc inferior vena cam.Blood smears ~v e i :~ niadc imniediatcly f o r each rat usi.rig 1 part of blood diluted with 3 p r t s of non-radi.oactive rat ser*um. 1.hc smears were a.ir dried, fixod f o r 4 niiniites in d)solutct methyl alcohol, arid stained for 1. hour with Wright ?s bl.ootl stain. Tlic wi(?xrs were t.ben coated wi.tl.1 unidlsioii €or mdioautography as indicated bclow and cxposed for 1' 2, 3 or 4 weeks.For fixat.io11, two pieccs of most, organs and tksues tverc r.cmoved and placed respec,tivcly i.n Rouin's fixative and. in 55: t.ri.chlorzlcetic acid. In order to insure obt.uining active hair follicles, skin samples extending from the dorm1 to the ventral midl.ine were taken from thc lower thoracic region. Tissues were kept in fixative for about a day wit,h the. cxccptioii of' head of tibia, lower jaw and tccth, foot pad, hppophysis, cerebrum and ccrcbellum, which were left in 5yj t,richloracct,ic acid for 3 weeks heforc processing.Sections were cut at 8 p and stained hy routine h,ematoxylineosin or kept unstained. Coated radioautographs (BQlaiiger and Lehlonci, '46 ; Gross ef al., '51) werc prepared by covering the scct.ions with melted KTR-3 emulsion (Eastman Kotlak) -Thc stained sets werc exposed for 10, 24 and 36 hours, the unstaincci set.s for 8,22, and 96 hours. r i

Studies of Protein Metabolism in Intestinal Epithelial Cells*

“…The heavy concentration of label in the proliferative cells of the crypts in jejunum and colon indicates rapid protein synthesis in the proliferating cells. With both leucine (8) and methionine (2), the labeling over proliferative cells is much heavier than over mature cells. The disappearance of label from the proliferative zones can be accounted for by emigration of labeled cells and continued protein synthesis from endogenous precursors.…”

Protein starvation and the small intestine

“…During normal nutrition the small intestine has a very rapid rate of protein synthesis (3,9,10,28), and this synthesis rate is not slowed markedly during protein depletion (3,11,21,24). After prolonged protein starvation parenterally administered radioactive amino acids are incorporated into the protein of the small intestinal mucosa to a greater extent than in any other organ (8,11,21).…”

“…In the earlier studies of the effect of protein depletion upon the incorporation of radioactive amino acids into intestinal mucosal protein, the amino acids were always given intraperitoneally (3,8,10,11,21) and it was found that there was more incorporation of the amino acid into small intestinal protein than in other tissues, and more than in the protein-fed animals. In the present study, oral administration of the amnino acid showed an even greater difference between protein-deprived and control animals.…”

“…The epithelium of the intestinal mucosa has a rapid turnover rate, which is reduced during protein deficiency (6) and starvation (7), but nevertheless this tissue synthesizes a large amount of protein each day (8,9). It is probably because of the rapid rate of growth that the small intestinal mucosa of the proteindeficient animal incorporates parenterally administered radioactive amino acids into protein more efficiently than other tissues (3,8,10,11).…”

Age changes in the canine pancreas: histomorphological, electron microscopic, and biochemical study

“…These techniques (Leblond et a^., 1957) are based on the fact that the first sites to become radioactive after injection of a labelled amino acid are those places where protein synthesis takes place. The radioactivity appearing later at other sites can be due to migration of the newly synthesized proteins (Warshawsky et , 1963).…”