Pre-M Phase-promoting Factor Associates with Annulate Lamellae in Xenopus Oocytes and Egg Extracts

We have used complementary biochemical and in vivo approaches to study the compartmentalization of M phase-promoting factor (MPF) in prophase Xenopus eggs and oocytes. We first examined the distribution of MPF (Cdc2/CyclinB2) and membranous organelles in high-speed extracts of Xenopus eggs made during mitotic prophase. These extracts were found to lack mitochondria, Golgi membranes, and most endoplasmic reticulum (ER) but to contain the bulk of the pre-MPF pool. This pre-MPF could be pelleted by further centrifugation along with components necessary to activate it. On activation, Cdc2/CyclinB2 moved into the soluble fraction. Electron microscopy and Western blot analysis showed that the pre-MPF pellet contained a specific ER subdomain comprising "annulate lamellae" (AL): stacked ER membranes highly enriched in nuclear pores. Colocalization of pre-MPF with AL was demonstrated by anti-CyclinB2 immunofluorescence in prophase oocytes, in which AL are positioned close to the vegetal surface. Green fluorescent protein-CyclinB2 expressed in oocytes also localized at AL. These data suggest that inactive MPF associates with nuclear envelope components just before activation. This association may explain why nuclei and centrosomes stimulate MPF activation and provide a mechanism for targeting of MPF to some of its key substrates

Analysis of the proteins synthesized in ultraviolet light-irradiated Escherichia coli following infection with the bacteriophages λ drif d 18 and λ dfus -3

The presence of EF-Tu, RNA polymerase subunit α, and EF-G on the λ dfus -3 genome and EF-Tu, ribosomal proteins L7/L12, and RNA polymerase subunit β on the λ drif d 18 genome has been confirmed using a two-dimensional gel electrophoresis technique sensitive to changes in isoelectric point and molecular weight. In this system two EF-Tu gene products could not be resolved. Following infection of ultraviolet light-irradiated Escherichia coli with either λ dfus -3 or λ drif d 18, the EF-Tu gene, tufA , near 65 minutes on the genetic map is expressed as 3–4 copies per EF-G molecule. The EF-Tu gene, tufB , near 79 minutes on the genetic map, is expressed at about one-third of this rate. α is expressed as 1 copy per EF-G molecule, β as 0.14 per EF-G molecule and L7/L12 as 2.5 per EF-G. These figures compare well with the relative amounts found in exponentially-growing cells, in which the ratio of EF-Tu to EF-G is approximately 5. Almost 90% of the total number of proteins (calculated on a molecular weight basis) which theoretically can be encoded on the λ drif d 18 have been identified on the two-dimensional gel.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/47541/1/438_2004_Article_BF00341733.pd

Chromosomal location of human genes encoding major heat-shock protein HSP70

The HSP70 family of heat-shock proteins constitutes the major proteins synthesized in response to elevated temperatures and other forms of stress. In eukaryotes members of the HSP70 family also include a protein similar if not identical to bovine brain uncoating ATPase and glucose-regulated proteins. An intriguing relation has been established between expression of heat-shock proteins and transformation in mammalian cells. Elevated levels of HSP70 are found in some transformed cell lines, and viral and cellular gene products that are capable of transforming cells in vitro can also stimulate transcription of HSP70 genes. To determine the organization of this complex multigene family in the human genome, we used complementary approaches: Southern analysis and protein gels of Chinese hamster-human somatic cell hybrids, and in situ hybridization to human chromosomes. We demonstrate that functional genes encoding HSP70 proteins map to human chromosomes 6, 14, 21, and at least one other chromosome .Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/45535/1/11188_2005_Article_BF01534692.pd

Cdks and the Drosophila cell cycle

Cyclin-dependent kinases play essential roles in driving the cell cycle. Much progress has been made in Drosophila over the past year in identifying the specific requirements for individual cyclins in particular cell cycle events. These studies encompass many aspects of the cell cycle, from the addition of a G(1) phase to the cell cycle during embryogenesis to the role of cyclin degradation in progression through anaphase