MONITORING OF PEROXISOME INDUCTION AND DEGRADATION BY FLOW CYTOMETRICANALYSIS OF HANSENULA-POLYMORPHA CELLS GROWN IN METHANOL AND GLUCOSE MEDIA - CELL-VOLUME, REFRACTIVE-INDEX AND FITC RETENTION
C. Smeraldi et al., MONITORING OF PEROXISOME INDUCTION AND DEGRADATION BY FLOW CYTOMETRICANALYSIS OF HANSENULA-POLYMORPHA CELLS GROWN IN METHANOL AND GLUCOSE MEDIA - CELL-VOLUME, REFRACTIVE-INDEX AND FITC RETENTION, Microbiology, 140, 1994, pp. 3161-3166
Cell refractive index has been used to monitor peroxisome behaviour in
the yeast Hansenula polymorpha by means of flow cytometry. Peroxisome
s are inducible organelles which may occupy a large fraction of the ce
ll volume when yeast cells are growing in methanol media. These organe
lles harbour a catalase that decomposes the hydrogen peroxide produced
in methanol oxidation by alcohol oxidase, a peroxisomal enzyme whose
subunits are arranged to form a regular crystalloid. Peroxisomes under
go a degradation process mediated by vacuoles whenever they and their
enzymes become metabolically redundant (e.g. during growth on glucose)
. Flow cytometric analyses of side scattered light (depending on cell
volume, morphology and structure) and fluorescein isothiocyanate reten
tion (due to the vacuole) were made on two wild-type strains of H. pol
ymorpha during exponential growth in glucose and methanol media and du
ring nutritional shifts from one carbon source to the other. The same
parameters were also analysed for a mutant strain only partially repre
ssed by glucose. We show that both the parameters are substrate-depend
ent and appear to reflect peroxisome development in the cells. The dat
a reported correlate well with the known cytological and biochemical d
ata, showing the possibility of using flow cytometry, a fast and sensi
tive technique, to analyse the dynamics of peroxisome proliferation an
d degradation in response to environmental as well as genetic factors.