Reversible phosphorylation of proteins in proliferating and differentiating cells : cyclic variations and the effect of growth regulators
Date
1994
Authors
Ferreira, Gracinda Maria Nunes
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Abstract
Living cells are highly auto-dynamic entities which
means that the underlying biochemistry is equally
dynamic, a reality which is ignored by most researchers.
Theoretical studies indicate that such a state must be
due to the existence of oscillatory variations in the
levels and activities of key components in the cell. In
this study, the dynamic behaviour of four major,
interrelated areas of cell biochemistry
(phosphorylation, dephosphorylation, the terminal
reaction of glycolysis and the amount of soluble
protein) were examined and all systems found to
oscillate in murine erythroleukaemic cells (MEL) and,
where examined, also in the human HL-6Q leukaemic cell
line.
certain processes have been shown to be oscillatory for the first time ( phosphorylation
potential, the lactate dehydrogenase active isozyme level and aspects of the regulation thereof). While others have been shown to occur at a higher frequency
than previously reported (phosphotyrosine phosphatase
activity, the activity and apparent isozyme pattern of
lactate dehydrogenase, the amount of extractable
protein). All rhythms are shown (for the first time) to
be complex and to involve several contributing
periodicities, some modulating the period and amplitude
of the observed oscillation. The frequencies are very
high (periods of 1-20 minutes and probably Less) and the
amplitudes are equally high (variations in magnitude of
as much as a hundred fold).
Phosphorylation processes, currently of particular
interest with regard to the nature and control of cell
proliferation are thus found to be more highly dynamic
than previously believed, a fact which throws some doubt
on the current ideas on cell proliferation. The actual
lactate dehydrogenase (LDH) active isozyme pattern is
shown not to be constant (as generally believed) but to
vary at high frequency (possibly due to
phosphorylation of the the enzyme) while the kinetics
and specificity of the lone isozyme in murine
erythroleukaemic cells appear to be varying at
equally high frequency due to the action of
regulators (perhaps arising elsewhere within the
glycolytic pathway). Similar results were obtained with
HL-60 leukaemic cells with at least two of the
isozymes varying in level, to some extent independently.
The hormone, insulin, and the inducer of cell
differentiation, HMBA (hexamethylenebisacetamide), have
been found to affect the dynamics of the four systems
although, because of the complexity of the rhythms the
actual effects on the dynamics are not easily defined.
Insulin has a marked effect on the mean level of
the activity of the LDH isozyme.
The fact that all oscillations are seen despite no
attempt being made to synchronise the cell population
suggests the existence of communication between cells
but how this can occur when the rhythms are of such high
frequency is intriguing.
All the results add further support for the long
standing view of my supervisor, that the properties and
behaviour of cells reflect the internal dynamics and
that differentiation, cancer and intracellular
signalling occur through changes in the pattern of
temporal organisation of cellular oscillations.
Description
A Dissertation Submitted to the Faculty of Science
University of the Witwatersrand, Johannesburg
In fulfilment of the requirements
for the Degree of Doctor of Philosophy
Keywords
Protein Kinases, Phosphorylation, Growth regulators