Dolichyl phosphate linked sugars as intermediates in the synthesis of yeast mannoproteins: an in vivo study

Freshly prepared protoplasts of Saccharomyces cerevisiae X 2180 incorporate [³H]mannose and [¹⁴C]glucose for about 30 min into glycolipids and mannoproteins. Among the radioactive glycolipids formed dolichyl phosphate mannose, dolichyl phosphate glucose and dolichyl pyrophosphate oligosaccharides have been identified. The oligosaccharides released by weak acid from the dolichyl pyrophosphate were treated with endo-N-acetylglucosaminidase H and separated by gel filtration on Bio-Gel P-4. The largest oligosaccharide obtained corresponded exactly in size to Glc₃Man₉GlcNAc₁ the compound formed also in animal tissues. Other oligosaccharides released from dolichyl pyrophosphate in addition to the glucose containing ones were mainly Man₉GlcNAc₁ and Man₈GlcNAc₁. No mannosyl oligosaccharide corresponding in size to the total inner core region found in native mannoproteins could be detected in a lipid-bound form.The radioactive dolichyl pyrophosphate oligosaccharides were formed transiently; after 40 min only about 40% of the maximal radioactivity was observed in this fraction. In the presence of cycloheximide this decrease did not take place.It is concluded that the dolichol pathway of N-glycosylation of glycoproteins in yeast cells is very similar, if not identical, to the reaction sequence worked out for animal cells.Dedicated to Professor Dr. Otto Kandler on his 60th birthday     

Ferritin as an exogenous yolk protein in snails

Summary A main yolk component in the oocytes of the pulmonate snailPlanorbarius corneus L. has been isolated and identified as the iron storage protein ferritin by its ultrastructure, iron content, immuunological properties and behaviour in disc electrophoresis. As judged from acrylamide electrophoresis data and ultrastructural observations, yolk ferritin is an exogenous protein which is synthesised in the hepatopancreas and taken up by the oocytes by endocytosis.     

Relationship between the chromatoid body and the acrosomal system in early spermatids of Myxine glutinosa L.

Summary A transient close relationship between the chromatoid body and the developing acrosome is demonstrated in early spermatids of Myxine glutinosa.This work was supported by the Norwegian Research Council for Science and Humanities (NAVF, Grant Nr. D 61.44) and the Austrian Fonds zur Förderung der wissenschaftlichen Forschung, Projekt 2183     

Teratoma formation by human embryonic stem cells: Evaluation of essential parameters for future safety studies

Transplantation of human embryonic stem cells (hESC) into immune-deficient mice leads to the formation of differentiated tumors comprising all three germ layers, resembling spontaneous human teratomas. Teratoma assays are considered the gold standard for demonstrating differentiation potential of pluripotent hESC and hold promise as a standard for assessing safety among hESC-derived cell populations intended for therapeutic applications. We tested the potency of teratoma formation in seven anatomical transplantation locations (kidney capsule, muscle, subcutaneous space, peritoneal cavity, testis, liver, epididymal fat pad) in SCID mice with and without addition of Matrigel, and found that intramuscular teratoma formation was the most experimentally convenient, reproducible, and quantifiable. In the same experimental setting, we compared undifferentiated hESC and differentiated populations enriched for either beating cardiomyocytes or definitive endoderm derivatives (insulin-secreting beta cells), and showed that all cell preparations rapidly formed teratomas with varying percentages of mesoderm, ectoderm, and endoderm. In limiting dilution experiments, we found that as little as two hESC colonies spiked into feeder fibroblasts produced a teratoma, while a more rigorous single-cell titration achieved a detection limit of 1/4000. In summary, we established core parameters essential for facilitating safety profiling of hESC-derived products for future therapeutic applications.