Sporophore production ofAgaricus bisporus in aseptic environments

Production of mature sporophores ofAgaricus bisporus was achieved for the first time in amended, autoclaved soil, gamma-sterilized soil, and soil-extract agar medium. The initiation of sporophores was triggered by metabolites of soil-inhabiting bacteria, particularly nodule forming isolates. Whether a single metabolite or several metabolites of these bacteria caused formation of sporophores could not be established; however, biotin alone when added to soil extract medium produced comparable results. The potentiality of different bacteria to induce sporophore formation varied considerably within species and isolates.Amino acids favored vegetative growth ofA. bisporus, but failed to induce formation of sporophores. Organic acids supported luxuriant growth and poor sporophore formation. Among several growth-promoting substances and vitamins, biotin induced abundant formation of mature sporophores.The authors are thankful to Dr. C. Corke, Department of Soil Microbiology, University of Guelph, Ontario, for providing some bacterial cultures used in this study.     

The essential role of hypusine in eukaryotic translation initiation factor 4D (eIF-4D). Purification of eIF-4D and its precursors and comparison of their activities

Eukaryotic translation initiation factor 4D (eIF-4D) is the only protein known to contain the amino acid, hypusine [N epsilon-(4-amino-2-hydroxybutyl)lysine]. This unusual amino acid is formed post-translationally by modification of a single specific lysine residue in an eIF-4D precursor protein. Two separate eIF-4D precursors, each of which contains a lysine residue in place of the hypusine residue and each of which thereby serves as a protein substrate for the hypusine modification, were purified from DL-2-difluoromethylornithine-treated Chinese hamster ovary cells by means of a five-step procedure. These two precursors termed PI and PII both have apparent molecular masses of approximately 17 kDa, indistinguishable from that of eIF-4D, but exhibit more acidic isoelectric points (5.1 and 5.25 for PI and PII, respectively, compared with 5.37 for eIF-4D). These physical characteristics, together with other properties, indicate that eIF-4D differs from PII only in possessing the hypusine residue in place of a lysine residue, whereas an additional structural difference exists between PI and eIF-4D. eIF-4D from CHO cells provides a significant enhancement of methionyl-puromycin synthesis, a model assay for translation initiation. Neither PI nor PII stimulates this in vitro system. These findings are the first direct evidence that hypusine is essential for the biological activity of eIF-4D.     

Label-retaining cells reside in the bulge area of pilosebaceous unit: implications for follicular stem cells, hair cycle, and skin carcinogenesis

Inconsistent with the view that hair follicle stem cells reside in the matrix area of the hair bulb, we found that label-retaining cells exist exclusively in the bulge area of the mouse hair follicle. The bulge consists of a subpopulation of outer root sheath cells located in the midportion of the follicle at the arrector pili muscle attachment site. Keratinocytes in the bulge area are relatively undifferentiated ultrastructurally. They are normally slow cycling, but can be stimulated to proliferate transiently by TPA. Located in a well-protected and nourished environment, these cells mark the lower end of the "permanent" portion of the follicle. Our findings, plus a reevaluation of the literature, suggest that follicular stem cells reside in the bulge region, instead of the lower bulb. This new view provides insights into hair cycle control and the possible involvement of hair follicle stem cells in skin carcinogenesis.     

A single tryptophan on M2 of glutamate receptor channels confers high permeability to divalent cations.

Ionotropic glutamate receptors (iGluRs) of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate/kainate subtype display lower permeability to Ca2+ than the N-methyl-D-aspartate (NMDA) subtype. The well-documented N/Q/R site on the M2 transmembrane segment (M2) is an important determinant of the distinct Ca2+ permeability exhibited by members of the non-NMDA receptor subfamily. This site, however, does not completely account for the different permeation properties displayed by non-NMDA and NMDA receptors, suggesting the involvement of other molecular determinants. We have identified additional molecular elements on M2 of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate/kainate receptor GluR1 that specify its permeation properties. Higher permeability to divalent over monovalent cations is conferred on GluR1 by a tryptophan at position 577, whereas blockade by external divalent cations is imparted by an asparagine at position 582. Hence, the permeation properties of ionotropic glutamate receptors appear to be primarily specified by two distinct determinants on M2, the well-known N/Q/R site and the newly identified L/W site. These findings substantiate the notion that M2 is a structural component of the pore lining.     

Evidence for ion chain mechanism of the nonlinear charge transport of hydrophobic ions across lipid bilayers.

The conductivity across a lipid bilayer by tetraphenylborate anion is increased 10-fold on the photoformation of lipophilic porphyrin cations. The cations alone have negligible conductivity. This nonlinear photogenerated increase of ion conductivity is termed the photogating effect. Substitution of H by Cl in the para position of tetraphenylborate leads to a 100-fold enhancement of conductivity, whereas the dark conductivities for this and other substituted borates are the same. Moreover, the halo-substituted borates show a large enhancement of conductivity in the low concentration range (10(-8) M), whereas that of tetraphenylborate is small and space charge is negligible. The enhanced ion conductivity has great structural sensitivity to the structure of the anion, the cation, and the lipid, whereas the partition coefficient of all the borates and the concentration of photoformed cations are only slightly affected. The photogated ion transport has a twofold larger activation energy than transport in the dark. Time-resolved photocurrents and voltages demonstrate that the translocation rate of the porphyrin cation is also enhanced 100-fold by the Cl-borate anion but only 10-fold by the H-borate anion. For these reasons the nonlinear gating effect cannot be explained by electrostatics alone, but requires an ion chain or ion aggregate mechanism. Kinetic modeling of the photoinduced current with a mixed cation-anion ion chain can fit the data well. The photogating effect allows the direct study of ion interactions within the bilayer.