Initiation of meiosis in cell cycle initiation mutants of Saccharomyces cerevisiae

Control of the initiation of meiosis in yeast was examined in diploids homozygous for one of four different temperature-sensitive mutations that affect “start” of the mitotic cell cycle. Two of the mutations, cdc28 and tra3, bring about deficiencies in the initiation of meiosis, while cdc25 and cdc35 do not prevent initiation of normal meiosis at both permissive and restrictive temperatures. Moreover, diploids homozygous for the latter two mutations are capable of initiating meiosis in rich growth media upon transfer to the high, non-permissive temperature. This unique feature contrasts with the behavior of other yeast strains which require a starvation sporulation medium for initiation of meiosis. It is suggested that the initiation of meiosis includes functions that are shared with “start” of the mitotic cell cycle, as well as functions related to the choice between the two processes. Meiosis in vegetative media at the restrictive temperature (in cdc25 or cdc35 homozygotes) may be important for the study of chemical and physiological phenomena resulting from the meiotic process and not from adaptation to the sporulation medium.     

Polarized Secretion of Drosophila EGFR Ligand from Photoreceptor Neurons Is Controlled by ER Localization of the Ligand-Processing Machinery

The release of signaling molecules from neurons must be regulated, to accommodate their highly polarized structure. In the developing Drosophila visual system, photoreceptor neurons secrete the epidermal growth factor receptor ligand Spitz (Spi) from their cell bodies, as well as from their axonal termini. Here we show that subcellular localization of Rhomboid proteases, which process Spi, determines the site of Spi release from neurons. Endoplasmic reticulum (ER) localization of Rhomboid 3 is essential for its ability to promote Spi secretion from axons, but not from cell bodies. We demonstrate that the ER extends throughout photoreceptor axons, and show that this feature facilitates the trafficking of the Spi precursor, the ligand chaperone Star, and Rhomboid 3 to axonal termini. Following this trafficking step, secretion from the axons is regulated in a manner similar to secretion from cell bodies. These findings uncover a role for the ER in trafficking proteins from the neuronal cell body to axon terminus.     

Dissection of the faint little ball (flb) phenotype: determination of the development of the Drosophila central nervous system by early interactions in the ectoderm.

The complex embryonic phenotype of mutations in the faint little ball (flb) locus, encoding the Drosophila EGF receptor homolog (DER), was dissected by temperature shifts of a temperature-sensitive allele. We show that the phenotype can be resolved into at least five components, which are temporally and spatially distinct. Most notably, the central nervous system (CNS) phenotype is determined at two separate phases. A severe collapse results from early defects in the DER-expressing ectodermal cells from which neuroblasts and midline glial cells deaminate. We thus suggest that DER activity is crucial for interactions that occur in the ectoderm at an early stage, and determine the fate of neuronal and glial cell lineages. This finding explains how a severe CNS phenotype is generated in flb embryos, in spite of the absence of expression of the protein in neuronal cells. In a second phase, during germ band retraction, the flb function is required specifically in the three pairs of midline glial cells (MG). In the absence of a functional DER protein, these cells die or fail to differentiate correctly, resulting in a fused commissure phenotype.     

Dependence of Marine Bdellovibrios on Potassium, Calcium, and Magnesium Ions

Marine bdellovibrios show a specific requirement for K⁺, Ca²⁺, and Mg²⁺. Potassium is essential for high velocity and seems to be necessary for attachment of the free bdellovibrios. Calcium and magnesium are necessary for attachment and penetration. Magnesium also plays a role in maintaining the integrity of the bdelloplast. The adaptation of these bdellovibrios to the marine environment is manifested by their stringent cation requirements.     

Assessing the Secretory Capacity of Pancreatic Acinar Cells

Pancreatic acinar cells produce and secrete digestive enzymes. These cells are organized as a cluster which forms and shares a joint lumen. This work demonstrates how the secretory capacity of these cells can be assessed by culture of isolated acini. The setup is advantageous since isolated acini, which retain many characteristics of the intact exocrine pancreas can be manipulated and monitored more readily than in the whole animal. Proper isolation of pancreatic acini is a key requirement so that the ex vivo culture will represent the in vivo nature of the acini. The protocol demonstrates how to isolate intact acini from the mouse pancreas. Subsequently, two complementary methods for evaluating pancreatic secretion are presented. The amylase secretion assay serves as a global measure, while direct imaging of pancreatic secretion allows the characterization of secretion at a sub-cellular resolution. Collectively, the techniques presented here enable a broad spectrum of experiments to study exocrine secretion.