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By
Omar El Mawas
A thesis submitted to the Department of Biology in partial fulfillment of the requirements for
the degree of Master of Sciences in Biology
Faculty of Sciences
University of Balamand
December 2015
University of Balamand
Faculty of Sciences
Omar El Mawas
and have found that it is complete and satisfactory in all the respects,
and that any and all revisions required by the final
examining committee have been made.
JURY MEMBERS:
Approved: ____________________
Full name, Ph.D.
President of the jury
Approved: ____________________
Full name, Ph.D.
Jury Member
Approved: ____________________
Full name, Ph.D.
Supervisor
ACKNOWLEDGEMENTS
I would like to express my sincere gratitude to my advisor Dr. .…… for his continuous
support, for his patience, motivation, enthusiasm, and immense knowledge. His guidance
helped me in all the time of research and writing of this thesis.
I also would like to thank my family for supporting me spiritually throughout my life.
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ABSTRACT
TABLE OF CONTENTS
ACKNOWLEDGEMENTS iii
ABSTRACT iv
TABLE OF CONTENTS v
LIST OF ABBREVIATIONS viii
LIST OF TABLES x
LIST OF FIGURES xi
CHAPTER 1: INTRODUCTION 1
1.1 The Cell and Cell Cycle 1
1.1.1 Cell Discovery 1
1.1.2 Cell Cycle: Description and Regulation 2
1.2 Cancer 6
1.2.1 Cancer Hallmarks 7
1.2.1.1 Limitless replication potential 7
1.2.1.2 Sustaining proliferative potential 8
1.2.1.3 Insensitivity to antigrowth signals 8
1.2.2 Proto-Oncogenes and Tumor Suppressor Genes 12
1.2.3 Cancer Risk Factors 13
1.2.3.1 Genetic predisposition 13
1.2.4 Cancer Sign and Symptoms 16
1.2.4.1 Unexpected weight loss 16
1.3.6.2.5 Inability to recover stalled forks 39
1.4 Calyculin A 40
1.5 Fluorescent in Situ Hybridization 42
CHAPTER 2: MATERIALS AND METHODS 49
2.1 Lymphocyte Culture and CFS Induction 49
2.2 Metaphase Spreading 49
CHAPTER 3: RESULTS 54
3.1 Lymphocyte Culture: Induction of Premature Chromosome Condensation and
54
Metaphase Spreading
vi
LIST OF ABBREVIATIONS
LIST OF TABLES
LIST OF FIGURES
CHAPTER 1
INTRODUCTION
The mid-17th century witnessed one of the possibly greatest scientific breakthroughs,
the discovery of the cell. Thanks to the advances in microscopy, English physicist Robert
Hooke was able to examine thin slices of cork in 1665. He noticed that such tissues are
formed by tiny pores, he termed cells from the Latin word Cella meaning 'a small room'. …
Further studies into the cell led Walther Flemming to the discovery that cells undergo cellular
division or Karyomitosis (meaning threadlike metamorphosis of the nucleus) and his work
was published in 1882. Cell division is required both in developmental and adult stages to
maintain homeostasis. In the adult stage, it replaces old, warn out cells with new functional
The order of events required through which a cell can pass from one cell division to the
next is termed cell cycle. Today, we know that the cell cycle consists of two successive
stages: interphase, a highly regulated long preparatory stage, and mitosis, a unidirectional and
tightly regulated process by which one parent cell divides to give rise to two identical
daughter cells. In most mammalian cells, Mitosis, lasting one hour, includes prophase,
metaphase, anaphase and telophase. While interphase, lasting around twenty-three hours,
includes G1, S and G2 phases. G1 is a preparatory growth stage for DNA replication and
division, S is the stage where replication takes place, and G2 is the stage where the cell
Most cells in an adult organism are not dividing. They remain inactive in a quiescent
stage called G0. Yet still each second twenty five million cells are undergoing cellular
such, specific cyclin proteins and cyclin-dependent kinases (CDKs) coordinately control the
progression of the cell from one stage to the next. CDKs are serine/threonine kinases.
Unbound CDKs are present in an inactive conformation (Collins & Garrett, 2005). The
them to become catalytically active. It is noteworthy that the levels of CDKs in the cell
remain moderately constant throughout the cell cycle. Apart from cyclin D levels, which
increases in the beginning of G1 and remains constant throughout, the levels of cyclin
fluctuate while the cell progresses through its cycle. Cyclins are classified according to the
stage where they are mostly elevated. The level of cyclin in the cell is determined by the rate
proteasome (Pecorino, 2012). The following Figure 1.1 highlights the different stages of the
cell cycle.
Growth factors such as c-MYC and c-fos induce the cell to leave G0 and re-enter the
cell cycle by inducing the expression of cyclin D and its partner CDK4/6. The interaction of
cyclin D with CDKs 4/6 leads the cell through G1…. Cyclin E interacts with CDK2 and
a conformational change takes place in the pocket domain of Rb causing the release of the
E2F. E2F target genes (cyclin A, thymidylate synthase, etc.) become fully expressed and the
cell can proceed through the S phase (Haering, Lowe, Hochwagen, & Nasmyth, 2002).
…Other CFS that have been shown to be associated with cancer include FRA7G which
is located at the band 7q31.2 and has been shown to be frequently expressed in prostate,
breast, and ovarian cancer (Huang, et al., 1998). Central deletions in FRA6E, where the
candidate tumor suppressor gene Parkin lies, have been shown to exist in ovarian and lung
cancers. The following Table 1.1 shows molecularly mapped CFS and Cancer associated CFS
Genes (CACGs).
The replication slippage model proposed by Sutherland et al. suggests that the Okazaki
fragment plays an important role during the mechanism of expansion of microsatellite CCG
or AT-rich minisatellites. In one case where the number of copies of repeats is less than 80, it
suggests that slippage of Okazaki fragment might still occur during polymerization though
the 5’ end of the Okazaki fragment might be firmly anchored by a unique sequence of DNA
at one side….
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