Genetics of Cancer

The incidences of cancer started shooting up in the year 1991. Since a decade the cancer cases increased by a factor of three. There is simply no complete cure to this dreadful condition. It involves changes in the genome leading to uncontrolled cellular proliferation, transformation, invasion, metastasis, apoptosis suppression, and angiogenesis. The environmental factors such as chemicals, radiations, viruses, microbes, and hormones cause cancer. Apart from the above reasons, there are more factors involved. The cell follows a cyclical pattern of division involving different phases. It includes the G1, S, G2, and the M phases respectively. The transition of one phase from the other involves checkpoints. The CDK/cyclin complexes mainly control the cell cycle. The checkpoints determine the damaged DNA. They also help in checking the problems in the cell cycle machinery. Hence, they play a crucial role in permitting normal cells to continue. Problems in the cell cycle checkpoints trigger cancerous conditions. Certain viruses such as retroviruses cause cancer. They increase the oncogenic products. Also, for the normal cells, apoptosis plays a crucial role. The cancer cells do not undergo apoptosis.

Image 1: Cancer cells and normal cells

Cell cycle and cancer:

A cell cycle involves six main checkpoints such as the restriction point, the G1/S DNA damage checkpoint, the S phase DNA damage checkpoint, G2/M checkpoint, centrosome duplication checkpoint, and mitotic checkpoint. The restriction point occurs between the mid to late G1 phase. This point ensures the cell to enter into the S phase after receiving the appropriate signals. The G1/S damage checkpoint occurs at the G1 phase transition. It senses the DNA damage. The S phase DNA damage checkpoint arrests the cell cycle in the later part. It detects the DNA damage or an incomplete replication of DNA. The G2/M checkpoint also detects the damaged DNA. The centrosome duplication checkpoint detects the defects in the centrosome duplication process. This checkpoint also detects centrosome segregation defects. The mitotic checkpoint occurs in the M phase. It checks the formation of mitotic spindles.

The CDK/cyclin complexes control the cell cycle. The cyclin-dependent kinases belong to the class of kinases. The cyclins are known as the regulatory subunits. The CDKs are known as catalytic subunits. The cell cycle checkpoints involve a genetic control. The genes participating in the multiple cell cycle checkpoints are known as gatekeeper genes. These genes prevent the cell cycle progression until the damaged DNA gets repaired.

Phases of the cell cycle	Cyclin-CDK complexes
G1 phase	Cyclin D, CDK-4 Cyclin D, CDK-6
Late G1 phase	Cyclin E, CDK-2
S phase	Cyclin A, CDK-2
G2 phase	Cyclin A, cdc 2
M phase	Cyclin B, cdc 2

Table: All phases of the cell cycle and the associated cyclin-CDK complexes

1.     G1 Phase:

Alterations in the signaling pathways associated with the cyclin-dependent kinases lead to the uncontrolled cell proliferation. Retinoblastoma involves tumor in the retina. It occurs in childhood. The gene responsible for getting mutated and causing the disease is known as the RB gene. It is present on the q arm of the thirteenth chromosome. Deletion or inactivation of both the copies of the RB gene leads to retinoblastoma. The cell loses the protein product pRb.

This phase requires a regulatory protein. It is known as pRb. It gets phosphorylated by cyclin/CDK complex. The pRb binds to the E2F transcription factor and prevents the cell’s entry into the S phase. After the phosphorylation of pRb, it gets inactivated and releases the E2F. Now the cell safely enters the S phase. However, in the retinoblastoma, the cell loses the pRb protein due to RB gene mutation. Hence, the cell enters into the S phase without checking any damaged DNA. Thus, it leads to an unrestrained tumor formation.

2.     G1/S checkpoint:

The tumor suppressor gene known as TP53 gene plays a crucial role in cell cycle arrest and DNA repair. This checkpoint gets invoked due to dsDNA breaks and damage. The product of the TP53 gene is a protein. It is known as p53. It helps in arresting the cell cycle in the G1 phase or the G1/S phase. After the repair of the DNA, the cycle resumes back. However, failure to get repaired leads to apoptosis or cell death. It occurs in the normal cells where p53 gets activated. In the cancer cells, the p53 is not present. Hence, there is no cell cycle arrest and repair of damaged DNA. Thus, the cells form tumors.

3.     G2/M checkpoint:

It is a DNA damage checkpoint. It helps in progressing the cell from the G2 phase to mitosis phase. It maintains the cdc2/ cyclin B1 in an inactive state. The protein p53 also plays a crucial role here.

Image 2: Cell cycle

Cellular proliferation:

Signal transduction involves extracellular growth factors. They regulate cell growth and differentiation. The genes encoding the growth factors or the growth factor receptors may get mutated. Hence, they lead to oncogenic properties. A gene encodes for the signal transducing protein. It is known as ras gene. The transcription factor gets encoded by another gene. It is known as the Myc gene. Mutations in both the genes also cause cancer.

Genes, Viruses, and Cancer:

Cancer involves mutations in three main gene classes. They include proto-oncogenes, tumor suppressor genes, and mutator genes. The products of proto-oncogenes stimulate cell proliferation. The mutant ones are known as oncogenes. They are the active forms of cancer genes. The oncogenes stimulate unregulated cellular proliferation. The RNA viruses also replicate via DNA intermediate. These viruses are known as retroviruses. Upon the retroviral infection, the RNA genome of the viral particle synthesizes a kind of cDNA. It is known as proviral DNA. The viruses also have oncogenes. They are known as viral oncogenes. When they occur in the host cell, these genes are known as cellular oncogenes. The host DNA sequences homologous to that of the virus are known as proto-oncogenes. These genes get activated to oncogenes. Three main methods do this. The first method involves increasing the amount of proto-oncogene product. The second method involves mutations in the coding sequences. Chromosomal translocation also leads to activation of oncogenes.

Apoptosis and Cancer:

The cell death or apoptosis gets triggered in the case of unrepaired damaged DNA or any other unwanted cellular conditions. The failure of the checkpoints in stopping the cell cycle progression also triggers cancer. Cancer also occurs due to the activation of anti-apoptotic genes such as Bcl_2. Thus, many such factors contribute to cancer. 

References:
[1] Human Genetics, 3/e, Gangane
[2] Molecular Genetics of Cancer, John Cowell
[3] The Genetics of Cancer: Genes Associated with Cancer Invasion, Metastasis, Gajanan V. Sherbet, M. S. Lakshmi
[4] API Textbook of Medicine, Ninth Edition, Two Volume Set, Y P Munjal, Surendra K Sharma

© Copyright, 2018 All Rights Reserved.

Eukaryotic cells undergo cyclical patterns of growth and division

The growth and developmental patterns observed in eukaryotic cells are different and complex as compared with prokaryotic cells. Also, the genetic material, the cellular compartmentalization, and mechanisms differ from that of prokaryotes. Eukaryotic cells are studied more than the prokaryotic cells because we humans are eukaryotes. The capacity of the cells to reproduce independently is tremendous. Hence, the study of the cell cycle helps to know exactly how a cell undergoes growth and division. The concept of two cells forming from a single cell gave rise to tissue and organ systems in the body. Thus it depends on how and when the cell undergoes a cyclical pattern. Every part of the cell has a specific role to play. Not only division but also the cells undergo differentiation. Thus there exist different types of cells such as immune cells, blood cells, sperm cells, and egg cells. The cells may be haploid or diploid containing one or two sets of chromosomes.

The genetic material is packed artistically in the cells. The chromosomes appear in different shapes when visualized under a microscope. We may describe a cell cycle as a process of cells in undergoing growth, replication of the genetic material and division. Before starting the discussion on the way cells undergo cyclical patterns, we must know the basic components of the cells. A eukaryotic cell structure consists of a nucleus and a cytoplasm. The nucleus is a house for the genetic material consists of DNA-protein interactive structures known as chromosomes. It is a place where DNA replication and transcription occurs. The extranuclear or outer part of the nucleus is known as the cytoplasm. It is a site for organelles and protein synthesis.

INFO BOX: Terminologies

1.     DNA: The genetic material in the nucleus. DNA is the blueprint of life. 2.     Chromosome: It is a linear structure composed of DNA-protein interaction. 3.    Chromatid: It is a subunit of the chromosome. The two chromatids constitute a chromosome. (Symmetrical halves running parallel to each other) 4.   Centromere: Chromatids are attached to each other through centromere. It is a constricted region. 5.     Centriole: It is a self-reproducing material involved in cellular processes. 6.  Nucleolus: It is RNA enriched spherical body associated with a chromosome segment.

Although a great variation exists in eukaryotic organisms and their cells, a typical eukaryotic somatic cell cycle duration is 24 hours. Four main stages of the cell cycle include the G₁ phase, S phase, G₂ phase, and M phase. The G1, S and G2 phases are clubbed and termed as interphase. The M phase or the mitotic phase is the division phase of the cell. The G1 phase is an initial period of growth or a pro synthetic gap with duration of ten hours. The S phase is the period of DNA synthesis. It lasts for nine hours. The G2 phase is a post-synthetic gap phase. It lasts for four hours. The duration of the mitotic phase is one hour. During the G1 phase, the chromosomes become thin and extended. The cell is responsive to growth signals. The G2 phase accompanies chromosome condensation. The cell cycle checkpoints regulate the transition from one phase to another. They monitor the integrity of the genome. The mitosis occurs in the somatic cells. Another process occurring in gametes is known as meiosis.

Mitosis:

The process of mitosis occurs in both haploid and diploid cells. Prophase, metaphase, anaphase, and telophase are stages of mitosis.

a.     Prophase: This is the initial stage of mitosis. It exhibits chromosome condensation and visibility. The spindle apparatus forms. A spindle is a set of tubulin fibers that move the eukaryotic chromosome during division. The spindle assembles outside the nucleus during the prophase. Early prophase, middle prophase, and late prophase are three subphases of prophase. In early prophase, the centrioles move apart. The chromosomes in early prophase reduce in their sizes. However, after some time they start getting visible. The nucleolus begins to disappear. The middle prophase involves the movement of the centrioles apart from each other. This phase is just a beginning of the mitotic spindle formation. The chromosomes are clearly visible in this phase. The late prophase involves the movement of centrioles towards the opposite sides. The spindle finally begins to form. The prophase chromosomes coil to produce a series of compact gyres. The kinetochore is a specialized protein that binds to the centromere. 

a.     Metaphase: It starts with the disappearance of the nuclear envelope. In this phase, the kinetochores are well attached to the centromeres so that the chromosomes align on the equator of the spindle. Hence their position is fixed on the equatorial plane. It is one plane halfway between the two spindle poles. The long axis of the chromosomes forms an angle of 90⁰ to the spindle axis. A metaphase plate is an equatorial plane to which the chromosomes are aligned. During metaphase, the alignment of the chromosomes is perfectly on the spindle. The processed cells get arrested in the metaphase for visualizing under an electron microscope. Visualization under an electron microscope reveals a dense framework of proteins (scaffold) surrounded by an uncoiled DNA. Hence it reveals a double-stranded DNA.

b.    Anaphase: Here, the centromeres of the sister chromatids separate forming two daughter chromosomes. The two chromatids separate by moving under the action of the traction fibers towards the spindle pole. Along with the chromatids, the kinetochores also separate. Due to disjunction, the sister chromatids get converted to independent chromosomes and move towards the poles by shortening the microtubules. Thus, the chromosomes acquire specific shapes such as V, J or rod-shaped. The position of the centromere decides the shapes of the chromosomes. A submetacentric chromosome is J-shaped. A metacentric chromosome is V-shaped. Anaphase is very crucial for the cell due to centromere splitting. However, incorrect centromere splitting could be catastrophic for the cell.

c.      Telophase: The grouping of the daughter chromosomes occurs in such a way that the chromosomes align in two groups at the opposite ends of the cell. The chromosomes begin uncoiling and form elongated shapes. The spindle disappears. Reconstruction of the nuclear envelope begins. The nucleoli reappear. The nuclear division completes at this point. The cells get two nuclei.

d.    Cytokinesis: Division of the cytoplasm and the compartmentalization of the two nuclei occur into separate cells.

Image 1: Phases of Mitosis

Meiosis:

It is a process occurring in the gametes. It results in the doubling of the gametic chromosomes resulting in the zygotic chromosome number. The process of meiosis involves a single chromosomal duplication followed by two successive nuclear divisions. It occurs after one DNA replication cycle. The original diploid nucleus undergoes two successive divisions.  It consists of one haploid set of chromosomes from the father and the other from the mother. Meiotic division and differentiation lead to the formation of the gametes (the sperm and the egg). In the first meiotic division, the chromosome number gets reduced from diploid to haploid. The second meiotic division is equivalent to mitotic division. Four stages of meiosis I include prophase I, metaphase I, anaphase I and telophase I.

a.     Prophase I: It is somewhat similar to mitotic prophase. However, the difference lies in the behavior of the chromosome and the crossing over process. Five subphases of the prophase I include leptotene, zygotene, pachytene, diplotene, and diakinesis. The coiling of chromosomes occurs in the leptotene stage. Through this phase, the cells begin its commitment to meiosis. Zygonema is the early mid-prophase I. In this phase the shortening of the chromosomes takes place. Chromosomes align themselves roughly and are known as homologous chromosomes. They are similar to each other and synapse during meiosis. The homologous chromosomes retain their genes from the common ancestors. The process of synapsis leads to the formation of zipper-like structures along the length of the chromatids. The synaptonemal complex aligns the two homologs precisely. The telomeres initiate the process of synapsis. They get clustered on the nuclear envelope forming a bouquet like arrangements. Telomeres assist the chromosomes to undergo synapsis by moving them around. The mid-prophase or pachynema stage starts after the completion of the synapsis. Since the homologous chromosomes consist of four chromatids, they are known as bivalents or tetrad. Here, the crossing over or a physical exchange takes place. If there are genetic differences in the homologous chromosomes, crossing over results into formation of new genetic material. This crucial step leads to genetic variation and recombination. It just involves reciprocal exchanges. At the end of the pachynema, the synaptonemal complex disassembles and enters into diplonema stage. The homologous chromosomes begin to move apart. The crossing over, as the name suggests, begins to look like a cross. It is known as chiasmata where the homologous chromosomes associate tightly. In the diakinesis, the nuclear envelope breaks down with spindle assembly.

b. Metaphase I: There is a complete breakdown of the nuclear envelope with the alignment of bivalents on an equatorial plane. Spindle formation and microtubule attachment take place.

c.  Anaphase I: The chromosomes disjoin and migrate to the opposite poles. Hence maternally and paternally derived centromeres move towards each pole. The segregated sister chromatids remain attached at their respective centromeres.

d. Telophase I: Formation of the new nuclear envelope occurs. The cell proceeds for cytokinesis.

Meiosis II with prophase II, metaphase II, anaphase II and telophase II results in the formation of four haploid cells from two haploid cells of meiosis I. The results of meiosis II in males are known as spermatids. They are haploids giving rise to spermatozoa or sperms. The result of meiosis II in females is called ootid and a polar body.

References:
[1] Medical genetics, G.P. Pal
[2] Human Genetics, 3/e, Gangane
[3] Vogel and Motulsky's Human Genetics: Problems and Approaches, Friedrich Vogel, Gunter Vogel, Arno G. Motulsky
[4] Biology for the IB Diploma: Standard and Higher Level, Andrew Allott
[5] Principles of Medical Genetics, Thomas D. Gelehrter

© Copyright, 2018 All Rights Reserved.

A review on human chromosomes

An Introduction to chromosomes:

The ability of the chromosomes in getting stained reveals the true nature of the chromosome (derived from the Greek work word. The word chroma indicates color). These stainable bodies appear like threads under a microscope. They contain the genetic material, mainly the DNA coiled around the proteins. Human chromosomes are visible with when the cell is undergoing mitotic or meiotic cell division. There are total 46 chromosomes in each human cell.

·    Autosomes: There are total 44 autosomes or 22 pairs. Each pair consists of homologous chromosomes. In a pair, one chromosome comes from the father and the other chromosome comes from the mother.

·        Sex Chromosomes: There are two different types of sex chromosomes, X and Y respectively.

The average size of the human metaphase chromosome is 5 millimeters. Chromosomes tightly coil and get condensed during metaphase. Chromosomes appear in different shapes during each phase of the cell cycle. They appear thread-like during interphase. During metaphase, chromosomes look like rod-shaped. They look like V, J or rod-shaped during anaphase. Von Hartz coined the term chromosome. The scientists who first discovered the structure of chromosomes were Schleiden, Virchow, and Bütschli. Walter Sutton and Theodor Boveri independently developed the chromosome theory of inheritance in 1902.

An interphase nucleus contains strands of a material called chromatin. There are two regions in the chromatin, mainly coiled and extended regions.

·    Heterochromatin: It is the dark staining area of the chromatin. There are two types of heterochromatin. Constitutive heterochromatin contains repetitive sequences. It is present near the centromere. Constitutive heterochromatin never expresses itself. There is one more type of heterochromatin, known as facultative heterochromatin, which expresses itself.

·        Euchromatin: It is the light staining area of the chromatin.

The chief constituent of chromatin is DNA, the blueprint of life. At the time of cell division, chromatin strands coil into compact structures, so that they easily fit into the cell nucleus. Chromosomes appear as thick rods only during the cell division. They uncoil and form chromatin at the end of cell division.

Image: Human chromosomes revealed through karyotyping

Structure of the chromosome:

Metaphase chromosome appears clearly under the microscope. Following are the principal point to be discussed:

·        Chromatids: Each metaphase chromosome consists of two symmetrical halves parallel to each other. They are called chromatids. These chromatids are present in the form of chromonema during prophase. There are two types of chromatids mainly, sister and non-sister chromatids. Two chromatids are present on a single chromosome. Thus they are called sister chromatids. The concept of dyad describes a pair of sister chromatids. These structures join with the help of a centromere.  Non-sister chromatids are either of the two chromatids of a chromosome pair.

·        Centromere: A centromere is a light staining constricted area to which both the chromatids are attached. A centromere divides the chromatids into short and long arms respectively. The short arm is known as “p” arm. The long arm is known as “q” arm. Centromere produces a primary constriction. It is the position of the centromere. It is different for different chromosomes. Secondary constriction is also known as nucleolar organizer region. It is involved in the formation of the nucleolus. Human centromere consists of several hundred kilobases of repetitive DNA. Centromeres are the sites where spindle fibers are attached. Thus, centromere helps in the movement of the chromosome.

·     Satellite: A satellite is a region that is attached to the chromosome by a thread of chromatin. It is present at the distal region of the arm of the chromosome.

·        Telomere: A special DNA-protein complex is present at the ends of the chromosomes. This complex is known as a telomere, with tandem repeats of TTAGGG-3’ sequences between 3-20 kilobases in length. Telomeres are not genes since they do not code for any functional molecule. Telomeres provide structural stability to the chromosomes by sealing their ends. Telomeres protect the chromosomes from damage. They also protect the chromosome from fusing into a ring or binding to other DNA.

Classification of chromosomes:

1.     Classification based on the position of the centromere:  

·     Metacentric: The two arms are almost equal in their lengths. The location of the centromere is at the center of the chromosome.

·   Submetacentric: The two arms of the submetacentric chromosomes are unequal in length. The location of the centromere is slightly away from the center.

·   Acrocentric: One arm of an acrocentric chromosome is short. Whereas, the other arm is long.

·      Telocentric: A telocentric chromosome has only one arm.

2.     Standard classification: It is also known as Denver classification. It classifies chromosomes into seven groups, depending on the length of the chromosomes.

·  Group A: This group consists of pairs of chromosomes 1, 2 and 3. Chromosome 1 is the largest human chromosome. It represents 8% of the total DNA content.  Chromosome 2 is the second one. The third chromosome represents 6.5 % of the total DNA content.

·       Group B: This group consists of pairs of chromosome 4 and 5.

·    Group C: This group consists of pairs of chromosome 6, 7, 8, 9, 10, 11 and 12.

·        Group D: This group consists of pairs of chromosomes 13, 14 and 15.

·        Group E: This group consists of pairs of chromosomes 16, 17 and 18.

·        Group F: This group consists of pairs of chromosomes 19 and 20.

·        Group G: This group consists of pairs of chromosomes 21 and 22.

·  Sex Chromosomes: There are two types of sex chromosomes. X chromosome and Y chromosome are called sex chromosomes.

3.     Paris nomenclature: According to this method, the long and short arms of the chromosomes have specific regions that get stained. These regions are further stained using banding techniques. Such techniques may not only help to identify specific chromosomes, but also find out the location within the chromosome. Banding techniques may help to detect minor structural abnormalities.

What is sex chromatin?

The nucleus in the interphase is in the resting phase. An interphase nucleus shows a dark stain chromatin mass attached on one side of the nuclear membrane. Sex chromatin is also known as the Barr body. It is observed only in females. However, the chromatin determination using the Barr body is not as accurate as the Karyotyping technique.

What are chromosomal aberrations?

Chromosomal mutations or aberrations are variation in the normal chromosome structure or chromosome number.

A deletion is a chromosomal mutation in which a part of a chromosome is missing. Chromosomal breaks result in deletions. Sometimes an entire chromosome may get deleted. Duplication may lead to doubling of a chromosomal segment. Excision of a chromosomal segment follows reinsertion leading to an inversion. A translocation is a chromosomal mutation in which a chromosome segment gets positioned in a different location in the genome.

Chromosome Analysis:

Chromosome analysis indicates a proper diagnosis of many clinical conditions. It is a microscopic analysis of chromosomes in the dividing cells. Chromosomal analysis can detect chromosome number and structure.

Uses of chromosome analysis are as follows:

·        Detection of congenital malformations, mental retardation, and repeated abortions.

·        Prenatal diagnosis

·        Diagnosis of malignancies

Karyotyping:

Karyotyping is a test to evaluate the number and the structure of the chromosomes. In this procedure, the metaphase chromosomes are obtained and photographed. The procedure of karyotyping is specialized. Peripheral blood lymphocytes, bone marrow cells or amniotic fluid samples are collected and analyzed for chromosomes. A photo-micrograph reveals chromosomes scattered randomly. These chromosomes are arranged into groups, using the software. A karyotype can be sued to detect chromosomal abnormalities.

Chromosome Banding:

Analysis of chromosome becomes precise with the help of banding techniques. There are four types of banding techniques such as G-banding, Q-banding, R-banding, and C-banding. A unique pattern of light and dark bands are obtained using the G-banding technique.

Fluorescence in-situ hybridization (FISH):

FISH is a new diagnostic technique that involves a single-stranded probe annealing to its complementary sequence. FISH can be used to detect minute chromosomal aberrations, malignancies, and study of chromosomes.

References:
[1] Medical genetics, G.P. Pal
[2] Human Genetics, 3/e, Gangane
[3] Vogel and Motulsky's Human Genetics: Problems and Approaches, Friedrich Vogel, Gunter Vogel, Arno G. Motulsky
[4] Biology for the IB Diploma: Standard and Higher Level, Andrew Allott
[5] Principles of Medical Genetics, Thomas D. Gelehrter

© Copyright, 2018  All Rights Reserved

Packaging of eukaryotic chromosomes

Eukaryotic species consist of a specific number of chromosomes. For example, humans have a total of 46 chromosomes. An average human chromosome consists of less than 5 cm of DNA. The DNA gets highly organized and packed into the chromosomes. Hence, the chromosomes are the most efficient genome packets.  The chromosomes have a different shape during every phase of the cell cycle.

However, the shape and the structure of the chromosomes get confusing in some species. The interphase and mitosis phase include the two phases of the cell cycle. The interphase consists of G1, S, and G2 phases. The chromosomes contain a thin and extended structure in the G1 phase. Hence, they do not appear clearly in this phase. The S phase is known as the phase of DNA replication. Hence, the chromosome gets duplicated by forming sister chromatids. The chromosome begins to condense in the G2 phase. The mitosis phase accompanies cell division. Hence, light microscopy helps to visualize the chromosomes in M phase. The metaphase chromosomes are thick and rod-shaped. However, the chromosomes in anaphase look like rods, J or V-shaped. Metaphase chromosomes form after DNA replication. The linking between the two chromatids involves a centromere. The chromosome arms are known as chromatids. A telomere is an extreme tip or the end of the chromosome. Metaphase chromosomes are present in a highly condensed form. 

Image: Packaging of eukaryotic chromosomes

Nucleosomes:

DNA packaging becomes difficult to study without understanding the nucleosome.  It is visible under an electron microscope. Each bead is known as a nucleosome. The string depicts the DNA. Each nucleosome consists of eight proteins known as histones. Five types of histones include H1, H2A, H2B, H3, and H4. Hence, each nucleosome consists of four main histones, each two in number. These proteins form a barrel-shaped core octamer with a DNA string wound twice around the barrel. The DNA linking the two nucleosomes is known as a linker DNA. The chromatin or a DNA histone complex gets stained with a suitable dye. The histones also contain large amounts of arginine and lysine. The charge of the amino acids is positive. Hence, it facilitates an easy binding with a negatively charged DNA. Like linker DNA, vertebrates consist of linker histones such as H1 a-e, H10, H1t, and H5. Each nucleosome gets attached to a linker histone acting like a clamp and preventing the coiled DNA from getting detached. Hence, the nucleosomes associate to form a 30nm fiber as revealed by the cell-breakage techniques. It involves interphase chromosomes in a highly condensed state or metaphase.

Non- histones are equally essential:

Non-histones prevail less abundantly as compared to the histones.  However, they play an equally important role. DNA binding proteins are known as histone proteins. They play an important role in DNA replication, repair, transcription, and recombination. They bind with the histones due to their negative charge. The proteins remaining in the chromatin after the removal of the histones are known as non-histones. Hence, the construction of the scaffold structure involves non-histone proteins. A scaffold structure is also known as the central framework of the chromosome. To this, a DNA solenoid gets attached as a loop(s). It also includes a class of enzymes known as topoisomerases. 

Two important models of a chromatin fiber:

1.   Solenoid model: A solenoid structure is known as a supercoiled arrangement of the DNA in the nuclear chromosome. This arrangement gets produced due to nucleosome string coiling. The process of coiling takes place continuously. It consists of 7 nucleosomes per turn.

2.     Helical ribbon model: It reveals the higher order structure of the chromatin.

Looped domains:

Selection micrographs show 30-90 kb loops of DNA. These loops get attached to a scaffold with a proteinaceous property. The X shape of the scaffold occurs due to the pairing of the sister chromatids. A human chromosome consists of 2000 looped domains. The location of the non-histone protein is at the base of the scaffold. Special regions get associated with the loops. They are known as scaffold-associated regions or SARs.

They involve a stretch of DNA binding to the non-histone proteins. The SARs determine the loops arranged spirally. The protrusion of the loops arises from the centromere.  They resemble petals of a flower with 15 loops per turn. Thus, the complex scaffolding helps in understanding the chromosomes in a better way.

The study of human chromosome:

Study of human chromosome involves cytogenetic technique such as karyotyping. Different banding techniques help in identifying the chromosomes. A typical metaphase chromosome in a human consists of a satellite, a centromere, a secondary constriction, and chromatids. The study of the sex chromatin and Barr body involves a separate study.

CEN sequences (DNA-protein interactions) in the yeast:

The centromere of the yeast has a single sequence with an approximately 125 base pairs length. This sequence consists of short regions known as CDE I and CDE III. A region known as CDEII lies between CDEI and CDEIII. CDEII is a longer element and is variable and AT-rich. The sequence of CDE I and CDE III are highly conserved. A histone H3 analog is known as Cse 4 protein. It is present in the yeast. This protein combines with another protein MIf2 and forms a core around which the CDEII sequence gets wrapped. Two more proteins get involved in the process. Cbf1 recognizes and attaches to the CDEI sequence. Cbf3 protein attaches to CDEIII. They altogether form the kinetochore. Unlike humans, yeast lacks nucleosomes. 

References:

[1] Cells: Molecules and Mechanisms, E.V. Wong

[2] Molecular Biology of the Gene, Watson

[3] Epigenetics, Lyle Armstrong

[4] Human Molecular Genetics, Tom Strachan

© Copyright, 2018 All Rights Reserved.