Search PubMed⌕ Search

Biomedical subjects

L Alberghina

Publications and source records attributed to L Alberghina.

99 records · Page 6Linked to original sources
PubMed1979

Intracellular protein degradation in Neurospora crassa.

In exponentially growing cultures of Neurospora crassa, the basal rate of protein degradation increases as the constant of the rate of growth decreases, so that in slow growing cells (mu = 0.13) the rate of protein degradation is about 25% of the rate of protein accumulation. During glucose starvation and shift-down transition of growth, the rate of protein degradation is greatly enhanced, and a moderate reduction (about 30%) of the ATP level is observed. Treatment of glucose-starved cells with 2-deoxyglucose reduces the ATP content by 70% and blocks protein degradation. The addition of cycloheximide, given at the onset of glucose starvation, prevents the enhancement of protein degradation; instead cycloheximide is without effect if added when proteolysis has already started. At a supraoptimal temperature (42 degrees C) the basal rate of protein degradation is not stimulated, contrary to the behavior observed in bacteria. Guanosine nucleotides, which appear to have a regulatory role for protein degradation in bacteria, are not found in N. crassa.

Amino Acids↗
PubMedDate not supplied

The sequence of 8.8 kb of yeast chromosome III cloned in lambda PM3270 contains an unusual long ORF (YCR601).

We have determined the nucleotide sequence of a segment of chromosome III contained in the right part of the lambda PM3270 clone, for a total of 8824 bp. This sequence contains an unusual long open reading frame, YCR601, of 6501 bp that encodes for a protein of 2167 amino acids that show no homology with other known proteins. YCR601 was disrupted by internal deletion and insertion of LEU2 gene and is a non-essential gene, however, it is transcribed during vegetative growth yielding a polyadenylated mRNA of approximately 7 kb.

Amino Acid Sequence↗
PubMedDate not supplied

A bimolecular mechanism for the cell size control of the cell cycle.

A molecular model for the control of cell size has been developed. It is based on two molecules, one (I) acts as an inhibitor of the entrance into S phase, and it is synthetised just after cell separation in a fixed amount per nucleus. The other (A) is an activator of the S phase, and it is synthetised at a ratio proportional to the overall protein accumulation. The activator reacts stoichiometrically with (I), and after all the (I) molecules have been titrated, (A) begins to accumulate. When it reaches a threshold value, it triggers the onset of DNA replication. This model was tested by simulation and when applied to the case of unequal division explains a number of features of an exponentially growing yeast cell population: (a) the lengths of TP (cycle time of parent cells) and TD (cycle time of daughter cells) verify the condition exp(- KTP ) + exp(- KTD ) = 1; (b) the changes of the average cell size of populations at different growth rates; (c) the frequency of parents and daughters at various growth rates; (d) the increase of cell size at bud initiation for cells of increasing genealogical age; (e) the existence of a TP - TB period (difference between the cycle time of parents and the length of budded phase) that depends linearly upon the doubling time of the population.

Cell Cycle↗
PubMedDate not supplied

Identification of different daughter and parent subpopulations in an asynchronously growing Saccharomyces cerevisiae population.

Under all growth conditions, a growing Saccharomyces cerevisiae yeast population is extremely heterogeneous, since individual cells differ in their cell size; this is due to their position in the cell division cycle and their genealogical age. To gain insight into the structure of a growing yeast population, we used a recently developed flow cytometric approach which enables, in asynchronously growing S. cerevisiae populations, tagging of both the cell age and the protein content of individual cells. This approach enabled the identification of daughter cells belonging to different cell cycle positions (i.e. newborn, G1, S + G2 + M + G1*, and dividing), thus yielding information about the relative fraction in the whole population, cell size and variability. More limited information could be obtained for the parent subpopulation; however, we were able to identify and characterize the dividing parent cells. The coefficient of variation (CV) of the protein content distribution for the dividing parents (27) was much higher than the CV of dividing daughters (18). Further findings obtained indicated a large overlap between the cell protein content distributions of daughter and parent cells as well as between the protein content of cells of the same subpopulation but belonging to different stages of the cell division cycle. The analysis of these differences enables a better understanding of the complex structure of an asynchronously growing yeast population.

Cell Cycle↗
PubMedDate not supplied

Flow cytometry and cell cycle kinetics in continuous and fed-batch fermentations of budding yeast.

Cell size distributions, obtained either as protein distribution by flow cytometry or as cell volume distribution by a Coulter counter, give relevant information about the growth conditions of populations of budding yeast Saccharomyces cerevisiae. We have previously found a good correlation between these distributions and the growth rate in continuous cultures (Ranzi et al., Biotechnol. Bioeng. 1986, 28, 185-190). We now present determinations of the protein distributions and cell volume distributions during different fed-batch fermentations performed with a simple on/off controller. Since during the fed-batch fermentation a true steady state is not obtained, the distributions continuously change with time, but nevertheless we observed a good correlation between the average of both distributions and the actual growth rate. The behavior of the cell size distributions can be interpreted on the basis of a two-threshold cell cycle model in which both the critical protein content at budding (Ps) and the critical protein content for cell division (Pm) are differently modulated by the growth rate. Additional findings will be presented showing that this model can be used to successfully explain the insurgence and the maintenance of oscillatory states in continuous cultures.

Cell Cycle↗
PubMedDate not supplied

Copy number modulation in an autoselection system for stable plasmid maintenance in Saccharomyces cerevisiae.

Efficient expression of a foreign gene requires a stable vector present at a high number of copies per cell. We have constructed an autoselection system for the stable maintenance of expression vector in the yeast Saccharomyces cerevisiae that uses the fructose 1,6-bisphosphate aldolase gene (FBA1) to stabilize plasmids in cells bearing a disruption of the chromosomal FBA1 gene. This system allowed us to obtain stable production of a reporter heterologous enzyme (Escherichia coli beta-galactosidase) in rich media. By using an inducible promoter to regulate the expression of FBA1 gene, we have also obtained the modulation of plasmid copy number by carbon source.

Bacterial Proteins↗
PubMedDate not supplied

Development of metabolically engineered Saccharomyces cerevisiae cells for the production of lactic acid.

Interesting challenges from metabolically engineered Saccharomyces cerevisiae cells arise from the opportunity to obtain yeast strains useful for the production of chemical(s). In this paper, we describe the accumulation of lactic acid in the culture medium of growing, engineered yeast cells expressing a mammalian lactate dehydrogenase gene (LDH-A). High and reproducible productions (20 g/L) and productivities (up to 11 g/L/h) of lactic acid have been obtained by modulating the physiological growth conditions. Since yeast cells are acid tolerant and survive at very low pH values, the production of lactate can be avoided. In perspective, the approaches described could be useful for the production of lactic acid, outflanking the problems related to the synthesis from bacteria cells. In fact, during industrial productions, there is an inhibitory effect on the metabolic activities of the growing bacteria (i.e., Lactobacillus spp.) caused by the acid produced and by the low pH value. Thus, strategies to prevent the lowering of pH are conventional operations. These processes allow the production of lactate(s) and require the purification of the acid from its salt. The biotechnological implications of this study are also discussed.

Animals↗
PubMedDate not supplied

Cell cycle and growth regulation in RAS2 mutant cells of Saccharomyces cerevisiae.

Yeast cells carrying ras2 temperature-sensitive mutations undergo a specific arrest in the prereplicative unbudded phase of the cell cycle when they are shifted to non-permissive temperatures. At 36.5 degrees C, in spite of their abnormally large cell size, bulk protein synthesis and accumulation rates are depressed in ras2 temperature-sensitive cells in comparison with isogenic wild type. At the same temperature, total RNA synthesis and accumulation rates are much more inhibited, suggesting that a defective Ras2/cAMP pathway alters the coordination between RNA and protein synthesis rates. The preferential RNA synthesis inhibition is correlated to a specific inhibition of the synthesis of the 35S rRNA precursor. These findings, taken together with the results of previous analyses, are in favour of a control by the cAMP pathway on rRNA biosynthesis.

Cell Cycle↗