Retroviral entry mediated by receptor priming and low pH triggering of an envelope glycoprotein.

Avian leukosis virus (ALV) has been used as a model system to understand the mechanism of pH-independent viral entry involving receptor-induced conformational changes in the viral envelope (Env) glycoprotein that lead to membrane fusion. Here, we report the unexpected finding that ALV entry depends on a critical low pH step that was overlooked when this virus was directly compared to the classical pH-dependent influenza A virus. In contrast to influenza A virus, receptor interaction plays an essential role in priming ALV Env for subsequent low pH triggering. Our results reveal a novel principle in viral entry, namely that receptor interaction can convert a pH-insensitive viral glycoprotein to a form that is responsive to low pH.

The Sec61p complex mediates the integration of a membrane protein by allowing lipid partitioning of the transmembrane domain.

We have investigated how the transmembrane (TM) domain of a membrane protein is cotranslationally integrated into the endoplasmic reticulum. We demonstrate that the Sec61p channel allows the TM domain to bypass the barrier posed by the polar head groups of the lipid bilayer and come into contact with the hydrophobic interior of the membrane. Together with the TRAM protein, Sec61p provides a site in the membrane, at the interface of channel and lipid, through which a TM domain can dynamically equilibrate between the lipid and aqueous phases, depending on the hydrophobicity of the TM domain and the length of the polypeptide segment tethering it to the ribosome. Our results suggest a unifying, lipid-partitioning model which can explain the general behavior of hydrophobic topogenic sequences.

Secreted cathepsin L generates endostatin from collagen XVIII.

Endostatin, an inhibitor of angiogenesis and tumor growth, was identified originally in conditioned media of murine hemangioendothelioma (EOMA) cells. N-terminal amino acid sequencing demonstrated that it corresponds to a fragment of basement membrane collagen XVIII. Here we report that cathepsin L is secreted by EOMA cells and is responsible for the generation of endostatin with the predicted N-terminus, while metalloproteases produce larger fragments in a parallel processing pathway. Efficient endostatin generation requires a moderately acidic pH similar to the pericellular milieu of tumors. The secretion of cathepsin L by a tumor cell line of endothelial origin suggests that this cathepsin may play a role in angiogenesis. We propose that cleavage within collagen XVIII's protease-sensitive region evolved to regulate excessive proteolysis in conditions of induced angiogenesis.

Signal sequence recognition in posttranslational protein transport across the yeast ER membrane.

We have analyzed how the signal sequence of prepro-alpha-factor is recognized during the first step of posttranslational protein transport into the yeast endoplasmic reticulum. Cross-linking studies indicate that the signal sequence interacts in a Kar2p- and ATP-independent reaction with Sec61p, the multispanning membrane component of the protein-conducting channel, by intercalation into transmembrane domains 2 and 7. While bound to Sec61p, the signal sequence forms a helix that is contacted on one side by Sec62p and Sec71p. The binding site is located at the interface of the protein channel and the lipid bilayer. Signal sequence recognition in cotranslational translocation in mammals appears to occur similarly. These results suggest a general mechanism by which the signal sequence could open the channel for polypeptide transport.

Signal sequence recognition in cotranslational translocation by protein components of the endoplasmic reticulum membrane.

We have investigated the role of membrane proteins and lipids during early phases of the cotranslational insertion of secretory proteins into the translocation channel of the endoplasmic reticulum (ER) membrane. We demonstrate that all steps, including the one during which signal sequence recognition occurs, can be reproduced with purified translocation components in detergent solution, in the absence of bulk lipids or a bilayer. Photocross-linking experiments with native membranes show that upon complete insertion into the channel signal sequences are both precisely positioned with respect to the protein components of the channel and contact lipids. Together, these results indicate that signal sequences are bound to a specific binding site at the interface between the channel and the surrounding lipids, and are recognized ultimately by protein-protein interactions. Our data also suggest that at least some signal sequences reach the binding site by transfer through the interior of the channel.

Molecular mechanism of membrane protein integration into the endoplasmic reticulum.

As proteins are integrated into the membrane of the endoplasmic reticulum, some hydrophilic polypeptide segments are transported through the translocation channel, others remain in the cytosol, and hydrophobic transmembrane sequences are released into the lipid phase. We have addressed the molecular mechanism by which these events occur. We demonstrate that both the lumenal and the cytosolic domains of a membrane protein are synthesized while the ribosome is membrane bound, so that even cytosolic domains come in contact with the translocation channel. We also find that, before translation of the protein is terminated, transmembrane sequences can laterally exit the translocation channel and enter the lipid environment. These results have significant implications for the folding and assembly of membrane proteins.

Sec61-mediated transfer of a membrane protein from the endoplasmic reticulum to the proteasome for destruction.

The human cytomegalovirus genome encodes proteins that trigger destruction of newly synthesized major histocompatibility complex (MHC) class I molecules. The human cytomegalovirus gene US2 specifies a product capable of dislocating MHC class I molecules from the endoplasmic reticulum to the cytosol and delivering them to the proteasome. This process involves the Sec61 complex, in what appears to be a reversal of the reaction by which it translocates nascent chains into the endoplasmic reticulum.

Systematic probing of the environment of a translocating secretory protein during translocation through the ER membrane.

We have extended a previously developed photo-crosslinking approach to systematically probe the protein environment of the secretory protein preprolactin, trapped during its transfer through the endoplasmic reticulum membrane. Single photoreactive groups were placed at various positions of nascent polypeptide chains of various length, corresponding to different stages of the transport process, and photo-crosslinks to membrane proteins were analyzed. In all cases, the polypeptide segment extending from the ribosome was found to be located in a membrane environment that is formed almost exclusively from Sec61 alpha, the multi-spanning subunit of the Sec61p complex that is essential for translocation. At early stages of the translocation process, before cleavage of the signal sequence, almost the entire nascent chain emerged from the ribosome contacts Sec61 alpha. The 'translocating chain-associating membrane' protein interacts mainly with the region of the signal sequence preceding its hydrophobic core. Our results suggest that the nascent chain is transferred directly from the ribosome into a protein-conducting channel, the major constituent of which is Sec61 alpha.

[Stomach perforation in the neonatal period and in children]. / Magenperforationen im Neugeborenen- und Kindesalter.

Among the upper gastrointestinal tract perforations in infants and children, the gastric perforations were predominating. With exception of peptic ulcers, localization was found mainly in the upper parts of the stomach regarding to mechanical originating factors. Mortality depends on prematurity and additional injuries as well as the basic disorders.

[An extragonadal retroperitoneal endodermal sinus tumor--results of treatment and treatment sequelae in a 3-year-old boy]. / Ein extragonadaler retroperitonealer Endodermal-Sinus-Tumor--Behandlungsergebnisse und Behandlungsfolgen bei einem 3 Jahre alten Knaben.

A case of an extragonadal endodermal sinus tumour in a three-year old boy is presented. Complete resection of the retroperitoneal tumour was possible after initial chemotherapy. Combined radio-chemotherapy after tumour removal led to complete remission now lasting for more than 5 years. There was unilateral renal atrophy and diverticulum of the stomach as treatment related complications. The favourable prognosis of this tumour due to intensified treatment is discussed in relation to treatment sequelae. The interdisciplinary approach in treatment of cancer in children is emphasized.

[Duplication of the intestinal tract]. / Duplikaturen des Verdauungstraktes.

Twelve duplications of the alimentary tract in eleven children are reviewed. There was a predominance of the spheric type of duplication as well as the neonatal group of children with clinical sign of tumour in most cases. There were no major problems in resection of spheric duplications but tubular type cases afforded different procedure in each case. Three cases with associated atresias were noted. There was one death related to gastric duplication.

[Complications caused by remnants of the omphalomesenteric duct]. / Komplikationen durch Rückbildungshemmung des Ductus omphaloentericus.

31 cases of complicated remnants of the omphalomesenteric duct are reviewed. In particular mechanical just as often as inflammatory complications are noted. There was a predominance of male infants with a mean age of 3.5 years. Surgery is recommended even for still non complicated Meckel's diverticula in childhood. There was no mortality.

[Varicocele in children and adolescents]. / Die Varikozele bei Kindern und Jugendlichen.

Idiopathic varicocele is quite common even among children and adolescents. Irreversible testicular damage may occur before adult age. Diagnosis and therapy for the purpose of preventing infertility constitute a challenge to youth health organisations and paediatric surgeons. Surgery according to Bernardt or Ivanissevich is considered the optimal therapeutic approach.

Follow-up studies in 50 totally colectomised children.

We report on 50 totally colectomised children, most of whom suffered from Hirschsprung's disease. Of the 50, one child died postoperatively of enteritis. On an average, the children were re-examined 5 1/2 years after the colectomy. The findings were as follows: With the exception of four, the size and weight of the patients were within the norm; 20 passed frequent stools of pulpy consistency; 16 suffered from disturbances of continence; 14 developed severe, partially recurrent enteritis. We did not find any advantage of a single method of operation, e.g. Martin's operation.

Investigations of trace analysis of A(III)B(V) semiconductor microsamples by atomic spectroscopy-VII Investigation of trace and thin-layer analysis of doping elements (Ag, Au, Bi, Cd, Sn, Tl) in InAs by atomic-absorption with electrothermal evaporation.

Etching procedures for separation of thin layers of InAs (up to 1.4 mum thick) have been developed and optimized. A solid-state microtome has been used for cutting layers thicker than 1mum. AAS/ETA methods for determination of traces of Ag, Au, Bi, Cd, Sn, and Tl have been developed. The matrix interferences of nitric add, hydrobromic acid, and As(3+) or In(3+) in nitric or hydrobromic acid have been studied. The main causes of the matrix interference are the formation of diatomic molecules between the trace and matrix components, and the effect of the evaporation processes. By use of the platform technique with special platforms some matrix interferences could be minimized. For the determination of traces of Sn, matrix modification with Ni(NO(3))(2) as additive gave the best analytical values. For the trace determination of Au, separation of In from Au by evaporation of InBr(3) from 0.6M HBR medium in the AAS ashing phase was developed. The detection limits are in the 10(16) atoms/cm(3) region for layers of 1 mum thickness and surface area 1 cm(2).

[Results of total colectomy in childhood]. / Folgen der totalen Colektomie im Kindesalter.

In nineteen colectomized children operated for Hirschsprung's disease in most instances, size and weight were within normal ranges about six years after surgery. They passed stools, mostly of pulpy consistence, once to ten-times a day, on average four times a day. Eleven children suffered from continence disturbances. Seven developed severe recurrent enteritis. These results as well as those of three children operated according to Martin's technique are compared with those in the literature.