Intracellular transport of mouse kidney β-glucuronidase induced by gonadotrophin

Abstract

1. The response of renal beta-glucuronidase with time to the injection of gonadotrophin was investigated in each submicrosomal fraction of rough and smooth microsomal fractions of mouse kidney homogenate. 2. The increase in beta-glucuronidase activity appeared initially in membranes of the rough microsomal fraction, 24h after injection. 3. Afterwards the newly synthesized enzyme appeared in the contents of the rough microsomal fraction and was subsequently found in the smooth microsomal fraction, reaching a maximum concentration in this fraction at 72h. 4. At this juncture, a decrease in the enzyme activity was observed in rough microsomal contents whereas the lysosomal fraction had reached its maximum value. 5. The time-course of the appearance of beta-glucuronidase in the submicrosomal fractions after the gonadotrophin stimulation suggests that the newly synthesized enzyme at the site of membrane-bound ribosomes is transferred across the membrane into cisternae of the rough endoplasmic reticulum, and then is transported into lysosomes via the smooth endoplasmic reticulum. 6. The properties of microsomal and lysosomal beta-glucuronidases were compared.

Physicochemical modifications of lysosomal hydrolases during intracellular transport

“…Evidence for the synthesis of lysosomal hydrolases in the microsomal fraction and their 1973 subsequent transfer to the lysosomal fraction has been presented for acid phosphatase and ,B-glucuronidase in rat liver van Lanckner & Lentz, 1970) and for ,8glucuronidase in mouse kidney (Ide & Fishman, 1969;Kato etal., 1970Kato etal., ,1972. Fishman and associates (Kato et al, 1972) have reported findings consistent with the view that ,-glucuronidase synthesized in gonadotrophin-stimulated mouse kidney is transported from the rough endoplasmic reticulum to the lysosomes via the smooth endoplasmic reticulum. A kinetic study of isotope incorporation into rat kidney subcellular fractions showed that labelled lysine, N-acetylglucosamine and mannose are incorporated into lysosomal peptides in a special rough microsomal fraction, whereas N-acetylneuraminic acid residues are incorporated into nascent glycoproteins in a smooth microsomal fraction enriched in Golgi membranes; labelled glycoproteins subsequently appear in lysosomes (Goldstone et al, 1971b;Goldstone & Koenig, 1972).…”

“…Golgi fractions from rat liver reportedly contain increased amounts of acid phosphatase activity (Cheetham et al, 1970;Fleischer & Fleischer, 1970) and arylsulphatase A and B activities (Nyquist, 1971). A smooth microsomal fraction containing Golgi membranes exhibited a higher acid phosphatase and ,B-glucuronidase activity than its rough counterpart from kidney of rat (Goldstone & Koenig, 1972) and mouse (Kato et al, 1972). These biochemical and cytochemical findings on Golgi fractions are consistent with previous electron-cytochemical observations in a wide variety of cell types showing the presence of acid phosphatase and arylsulphatase in situ in portions of the Golgi apparatus (Ericksson, 1969;Smith, 1969;Nichols et al, 1971).…”

“…The RNA content of the other fractions declined sharply in the direction of the SM1, GM and L fractions, whereas the phospholipid content declined more gradually in the same direction. It can be seen in Table 1 that the chemical composition of the RM1, SM and GM fractions from rat kidney approximated that of comparable fractions from mouse kidney (Kato et al, 1972) and rat liver (Dailner, 1963;Leelavathi et al, 1970).…”

“…Protein, RNA and phospholipid content of rat kidney subcellular fractions Subcellular fractions were prepared and analysed for protein, RNA and phospholipid as described in the text. The values for mouse kidney are from Kato et al (1972). The values for rat liver RNA and phospholipid are from Leelavathi et al (1970) and Dallner (1963) The RM2 fraction represented about 2% of the protein in the RM1 fraction and had the same phospholipid content but a slightly lower RNA content than the latter.…”

Rabbit β-glucuronidase. Subcellular distribution and immunochemical properties

“…Of a variety of treatments used, 0.5% sodium deoxycholate and 0.1 % octyl sodium sulphate were most effective, and both rendered 85-90% of the enzyme non-sedimentable. Deoxycholate solubilization of 8-glucuronidase has been used previously by Kato et al (1972), and octyl derivatives have been suggested as very effective alternatives (Tanford, 1972). Ribonuclease digestion, as used by Van Lancker & Lentz (1970), was not necessary.…”