Membrane traffic between secretory compartments is differentially affected during mitosis.

Abstract

Membrane traffic has been shown to be regulated during cell division. In particular, with the use of viral membrane proteins as markers, endoplasmic reticulum (ER)-to-Golgi transport in mitotic cells has been shown to be essentially blocked. However, the effect of mitosis on other steps in the secretory pathway is less clear, because an early block makes examination of following steps difficult. Here, we report studies on the functional characteristics of secretory pathways in mitotic mammalian tissue culture cells by the use of a variety of markers. Chinese hamster ovary cells were transfected with cDNAs encoding secretory proteins. Consistent with earlier results following viral membrane proteins, we found that the overall secretory pathway is nonfunctional in mitotic cells, and a major block to secretion is at the step between ER and Golgi: the overall rate of secretion of human growth hormone is reduced at least 10-fold in mitotic cells, and export of truncated vesicular stomatitis virus G protein from the ER is inhibited to about the same extent, as judged by acquisition of endoglycosidase H resistance. To ascertain the integrity of transport from the trans-Golgi to plasma membrane, we followed the secretion of sulfated glycosaminoglycan (GAG) chains, which are synthesized in the Golgi and thus are not subject to the earlier ER-to-Golgi block. GAG chains are valid markers for the pathway taken by constitutive secretory proteins; both protein secretion and GAG chain secretion are sensitive to treatment with n-ethyl-maleimide and monensin and are blocked at 19 degrees C. We found that the extent of GAG-chain secretion is not altered during mitosis, although the initial rate of secretion is reduced about twofold in mitotic compared with interphase cells. Thus, during mitosis, transport from the trans-Golgi to plasma membrane is much less hindered than ER-to-Golgi traffic. We conclude that transport steps are not affected to the same extent during mitosis.

In Vitro Synthesis of Sulfated Glycosaminoglycans Coupled to Inter-compartmental Golgi Transport

“…3B). This result agrees with an early study with CHO cells (54) in which a 3-4-fold inhibition of GAGs synthesis under mitosis was reported. The decrease in GAG synthesis could also be interpreted as the consequence of the effect of nocodazole per se on the GAG biosynthetic machinery or the result of widespread cell death after 24 h with the drug rather than a specific effect arising from the mitotic condition.…”

In vitro fusion of endocytic vesicles is inhibited by cyclin A-cdc2 kinase.

“…More likely, they may reflect a difference in pathways downstream from cyclin Acdc2 kinase in Xenopus and mammalian extracts or even differences between the way that membrane traffic is controlled during cell division in the two systems. Several such differences are apparent in vivo; endoplasmic reticulum-Golgi transport is blocked in mammalian cells (Featherstone et al, 1985) but not in maturing Xenopus oocytes (Ceriotti and Colman, 1989;Kanki and Newport, 1991), whereas the inverse is found for transGolgi-cell surface transport (Kreiner and Moore, 1990;Leaf et al, 1990). It is not known whether the endocytic pathway is blocked in maturing oocytes.…”

Okadaic Acid Induces Selective Arrest of Protein Transport in the Rough Endoplasmic Reticulum and Prevents Export into COPII-Coated Structures

“…The second line of evidence comes from studies of Golgi membrane traffic which have revealed that import predominates over export. Thus, import pathways from the rough ER (RER) and from the plasma membrane via endocytosis are significantly inhibited in both mitotic cells and OA-treated cells (12,13,24,35,67), while export pathways such as those carrying glycosaminoglycans and the TGN marker TGN 38 out of the TGN are much less affected and appear to be active during both mitosis (24) and OA treatment (22).…”