Biomedical subjects

Lelio Orci

Publications and source records attributed to Lelio Orci.

22 records · Page 2Linked to original sources

PubMed2002

Sec16p potentiates the action of COPII proteins to bud transport vesicles.

SEC16 encodes a 240-kD hydrophilic protein that is required for transport vesicle budding from the ER in Saccharomyces cerevisiae. Sec16p is tightly and peripherally bound to ER membranes, hence it is not one of the cytosolic proteins required to reconstitute transport vesicle budding in a cell-free reaction. However, Sec16p is removed from the membrane by salt washes, and using such membranes we have reconstituted a vesicle budding reaction dependent on the addition of COPII proteins and pure Sec16p. Although COPII vesicle budding is promoted by GTP or a nonhydrolyzable analogue, guanylimide diphosphate (GMP-PNP), Sec16p stimulation is dependent on GTP in the reaction. Details of coat protein assembly and Sec16p-stimulated vesicle budding were explored with synthetic liposomes composed of a mixture of lipids, including acidic phospholipids (major-minor mix), or a simple binary mixture of phosphatidylcholine (PC) and phosphatidylethanolamine (PE). Sec16p binds to major-minor mix liposomes and facilitates the recruitment of COPII proteins and vesicle budding in a reaction that is stimulated by Sar1p and GMP-PNP. Thin-section electron microscopy confirms a stimulation of budding profiles produced by incubation of liposomes with COPII and Sec16p. Whereas acidic phospholipids in the major-minor mix are required to recruit pure Sec16p to liposomes, PC/PE liposomes bind Sar1p-GTP, which stimulates the association of Sec16p and Sec23/24p. We propose that Sec16p nucleates a Sar1-GTP-dependent initiation of COPII assembly and serves to stabilize the coat to premature disassembly after Sar1p hydrolyzes GTP.

Biological Transport↗

PubMed2002

A resident Golgi protein is excluded from peri-Golgi vesicles in NRK cells.

To understand the structure and the function of the Golgi apparatus, it is essential to establish how resident Golgi enzymes are localized in only a few Golgi cisternae. In particular it is crucial to establish whether Golgi enzymes are retained specifically in cisternae, or if they are continuously transported from cisterna to cisterna. Here we report that a resident Golgi enzyme is largely excluded from peri-Golgi transport vesicles in normal rat kidney cells, a cell type in which conflicting results have been reported. Analysis of the lateral distribution of two markers within Golgi cisternae led to the same conclusion: a protein incorporated in vesicles (KDEL receptor) is concentrated at the rims of cisternae where vesicles form, while mannosidase II is not. These results suggest that localization of resident Golgi enzymes is achieved primarily by selective retention within cisternae and exclusion from transport vesicles. These observations cannot easily be reconciled with the vision of rapidly maturing Golgi cisternae as the principal means of intra-Golgi transport.

Animals↗

PubMed2002

Severe block in processing of proinsulin to insulin accompanied by elevation of des-64,65 proinsulin intermediates in islets of mice lacking prohormone convertase 1/3.

The neuroendocrine processing endoproteases PC2 and PC1/3 are expressed in the beta cells of the islets of Langerhans and participate in the processing of proinsulin to insulin and C-peptide. We have previously shown that disruption of PC2 (SPC2) expression significantly impairs proinsulin processing. Here we report that disruption of the expression of PC1/3 (SPC3) produces a much more severe block in proinsulin conversion. In nulls, pancreatic and circulating proinsulin-like components comprise 87% and 91%, respectively, of total insulin-related immunoreactivity. Heterozygotes also show a more than 2-fold elevation in proinsulin levels to approximately 12%. Immunocytochemical and ultrastructural studies of the beta cells reveal the nearly complete absence of mature insulin immunoreactivity and its replacement by that of proinsulin in abundant immature-appearing secretory granules. In contrast, alpha cell morphology and glucagon processing are normal, and there is also no defect in somatostatin-14 generation. Pulse-chase labeling studies confirm the existence of a major block in proinsulin processing in PC1/3 nulls with prolongation of half-times of conversion by 7- and 10-fold for proinsulins I and II, respectively. Lack of PC1/3 also results in increased levels of des-64,65 proinsulin intermediates generated by PC2, in contrast to PC2 nulls, in which des- 31,32 proinsulin intermediates predominate. These results confirm that PC1/3 plays a major role in processing proinsulin, but that its coordinated action with PC2 is necessary for the most efficient and complete processing of this prohormone.

Animals↗

PubMed2002

Microscopy: an art?

There can be no doubt that the finest creator of beauty is Mother Nature. And in many ways, science is the exploration of this beauty and of the mechanisms that have created it. Microscopy, as a technique in scientific investigation, has had a key role in uncovering nature's beauty, which has led some to propose that microscopy could be described as an art or even an art form. But is this claim justified?

Anatomy, Artistic↗