Pluripotency is the defining characteristic of human embryonic stem cells (hESCs) allowing them to differentiate into any somatic cell in the human body. For the promising clinical applications of hESCs, improved regulation of pluripotency and differentiation trajectories of their colonies is required . The pluripotency transcription factors (PTFs) which regulate pluripotency are inherently stochastic (with small fluctuations impacting cell fate), inherited asymmetrically upon cell divisions, and more similar in closely related (separated by less division events) cells. Here we use time-lapse experimental data of OCT4 fluorescence intensity to quantify the temporal  dynamics of the PTF OCT4 over a cell lifetime . We characterise the internal self-regulation of OCT4 using the Hurst exponent and autocorrelation analysis, quantify the intra-cellular fluctuations and consider the diffusive nature of OCT4 evolution for individual cells and pairs of their descendants. After cell divisions, OCT4 abundance in the daughter cells fluctuates sub-diffusively, showing anti-persistent self-regulation with a Hurst exponent of 0.38. Auto-correlation analysis shows anti-persistence for five hours or longer in 86\% of cells, on average between three and 12 hours into the cell cycle. The OCT4 fluctuations in five minute intervals follow a Laplace distribution, with BMP4 addition provoking smaller changes and tighter self-regulation, particularly in the differentiated fate group. This quantitative framework provides a basis for comparison to other experiments, and the development of mathematical models of pluripotency .
The improved in-vitro regulation of human embryonic stem cell (hESC) pluripotency and differentiation trajectories is required for their promising clinical applications. The temporal and spatial quantification of the molecular interactions controlling pluripotency is also necessary for the development of successful mathematical and computational models . Here we use time-lapse experimental data of OCT4-mCherry fluorescence intensity to quantify the temporal and spatial dynamics of the pluripotency transcription factor OCT4 in a growing hESC colony in the presence and absence of BMP4 . We characterise the internal self-regulation of OCT4 using the Hurst exponent and autocorrelation analysis, quantify the intra-cellular fluctuations and consider the diffusive nature of OCT4 evolution for individual cells and pairs of their descendants. We find that OCT4 abundance in the daughter cells fluctuates sub-diffusively, showing anti-persistent self-regulation . We obtain the stationary probability distributions governing hESC transitions amongst the different cell states and establish the times at which pro-fate cells (which later give rise to pluripotent or differentiated cells) cluster in the colony. By quantifying the similarities between the OCT4 expression amongst neighbouring cells, we show that hESCs express similar OCT4 to cells within their local neighbourhood within the first two days of the experiment and before BMP4 treatment. Our framework allows us to quantify the relevant properties of proliferating hESC colonies and the procedure is widely applicable to other transcription factors and cell populations .
R
Pluripotency is the defining characteristic
The improved in-vitro regulation
R
cells (hESCs) allowing them to differentiate into any somatic cell in the human body. For the promising clinical applications of hESCs, improved regulation of
cell (hESC)
R
of their colonies is required
is required for their promising clinical applications. The temporal and spatial quantification of the molecular interactions controlling pluripotency is also necessary for the development of successful mathematical and computational models
D
The pluripotency transcription factors (PTFs) which regulate pluripotency are inherently stochastic (with small fluctuations impacting cell fate), inherited asymmetrically upon cell divisions, and more similar in closely related (separated by less division events) cells.
R
OCT4
OCT4-mCherry
A
and spatial
R
PTF
pluripotency transcription factor
R
over a cell lifetime
in a growing hESC colony in the presence and absence of BMP4
R
After cell divisions,
We find that
R
with a Hurst exponent of 0.38. Auto-correlation analysis shows anti-persistence for five hours or longer in 86\% of cells, on average between three and 12 hours into the cell cycle. The
. We obtain the stationary probability distributions governing hESC transitions amongst the different cell states and establish the times at which pro-fate cells (which later give rise to pluripotent or differentiated cells) cluster in the colony. By quantifying the similarities between the
R
fluctuations in five minute intervals follow a Laplace distribution, with
expression amongst neighbouring cells, we show that hESCs express similar OCT4 to cells within their local neighbourhood within the first two days of the experiment and before
R
addition provoking smaller changes and tighter self-regulation, particularly in the differentiated fate group. This quantitative framework provides a basis for comparison to other experiments, and the development of mathematical models of pluripotency
treatment. Our framework allows us to quantify the relevant properties of proliferating hESC colonies and the procedure is widely applicable to other transcription factors and cell populations