GENE AND CHROMOSOMES

 CHROMOSOMES

transmission and expression of genetic information. a centromere which is most evident at metaphase where it is the narrowest part of the chromosome and the region at which spindle fibres attach. replication origins certain DNA sequences along each chromosome at which DNA replication is initiated. telomeres are the end of a linear chromosome that has a specialized structure to prevent internal DNA from being degraded by nucleases. 

  

BACTERIAL CHROMOSOME

appears as a distinct clump, the nucleoid , which is confined to a definite region of the cytoplasm.

if a bacterial cell is broken open gently, its DNA spills out in a secret of twisted loops. the ends of the loops are most likely held in place by protein. many bacteria contain additional DNA  6in the form of circular molecules called plasmids.

EUKARYOTIC CHROMOSOME

individual eukaryotic chromosomes contain enormous amount of DNA  and consists of a single, extremely long molecules of DNA. the chromosomes are in elongated, relatively uncondensed state  during interphase of the cell cycle. in the course of the cell cycle, the level of DNA packaging changes chromosomes progress from a highly packed state to a state of extreme condensation. DNA packaging also changes locally in replication and transcription when the two nucleotide strands must unwind so that particular base sequences are exposed. thus, the packaging of eukaryotic DNA is not static but changes regularly in response to cellular processes.

PACKAGING DNA INTO SMALL SPACES

chromosome of bacteria E. coli. a single molecule of DNA with approximately 4.64 million BP stretched out straight. DNA would be about 1000 times as long as the cell within which it resides. human cells contain 6 million base pairs of DNA which would measure some 1.8 m stretched end to end. even DNA in the smallest human chromosomes would stretch 14000 times the length of the nucleus. DNA molecules must be tightly packed to fit into such small spaces.

CHROMATIN STRUCTURE

eukaryotic DNA is closely associated with protein creating  chromatin. two basic types are:

1. euchromatin: which undergoes the normal process of condensation and condensation in the cell cycle. 

2. heterochromatin: which remains in a highly condensed state throughout the cell cycle, even during interphase.

it is of two types:

a. constitutive: always inactive and condensed. consists of repetitive DNA. late replicating and AT rich region . present at identical positions on all chromosomes in all cell types of an organism. genes poorly expressed . human chromosome 1,9,16, and Y chromosomes contain large region of constitutive heterochromatin. occur around the centromere and near telomere.

b. facultative: genetically active (decondensed) and active (condensed), variable in its expression. it varies with the cell types and may be manifested as condensed and heavily stained.

protein in chromatin are the histones , which are relatively small, positively charged protein of five major types : H1, H2A, H2B, H3, H4.

all histones have a high percentage of arginine and lysine positively charged amino acid  that give them a net positive charge. the positive charges attract the negative charges on the phosphate of DNA and holds the DNA in contact with the histones.

CHROMOSOME MORPHOLOGY

under the microscope chromosome appear as thin, thread like structures .

they all have a short arm and long arm separated by a primary constriction called ceteromere. the short arm is designated as p and long arm as q. the centromere. it is essential for the normal movement and segregation of chromosomes during cell division.

human metaphase chromosomes can be catogeried according to the length of the short and long arm and also the centromere location.

Metacentric: chromosomes have short and long arms of roughly equal  length with the centromere in the middle.

Submetacentric: chromosomes have short and long arms of unequal length with the centromere more towards one end.

Acrocentric: have a centromere very near to one end and have very small short arms. they frequently have secondary constriction on the short arms that connect very small pieces of DNA called stalk and satellites , to the centromere. 

the stalk contain genes which code for ribosomal RNA.. Ideograms are a schematic representation of chromosomes.

 

Each human chromosome has a 

short arm ("p" for "petit") and

 long arm ("q" for "queue"),

separated by a centromere. The ends of the chromosome are called telomeres.

Each chromosome arm is divided into regions, or cytogenetic bands, that can be seen using a microscope and special stains. 

The cytogenetic bands are labeled p1, p2, p3,   q1, q2, q3, etc., counting from the centromere out toward the telomeres. At higher resolutions, sub-bands can be seen within the bands. 

The sub-bands are also numbered from the centromere out toward the telomere.

CHROMATIN STRUCTURE

non histone chromosomal proteins make up about half of the protein mass of the chromosome. non- histone chromosomal proteins serves the structural roles and those that takes part in genetic processes such as transcription and replication. chromosomal scaffold proteins are revealed when chromatin is treated with concentrated salt  solution which removes histones and most other chromosomal protein, leaving a chromosomal protein skeleton to which the DNA is attached. these scaffold proteins may play a role in the folding and packaging of chromosome. other structural protein make up the kinetochore, cap the chromosome ends by attaching to telomeres, and constitute the molecular motors the move chromosomes in mitosis and meosis. other types of non histone chromosomal protein plays a role in genetic processes. components of the replication machinery. high mobility group proteins are small, highly charged proteins are small, highly charged proteins that vary in amount and composition, depending upon tissue type and stage of the cell cycle. these proteins may play an important role in altering the packaging of chromatin during transcription.

NUCLEOSOME

chromatin has a highly complex structure with several levels of organisation. the simplest level is the double helical structure of DNA. at a more complex level,the DNA molecules is associated  with proteins and is highly folded to produce a chromosome. when chromatin is isolated from the nucleus of a cell and viewed with an electron microscope, it frequently looks like beads on string. if a small amount of nuclease is added to this structure, the enzyme cleaves the string between the beads, leaving individual beads attached to about 200 bp of DNA. the repeating core of protein and DNA produced by digestion with nuclease enzymes is the simplest level of chromatin structure called nucleosome. the nucleosome is a core particle consisting of DNA wrapped about two times around an octamer of eight histone proteins (two copies each of H2A,H2B, H3, H4). the DNA in direct contact with the histone octamer is between 145 and 147 bp in length, coils around the histones in the left handed direction and is super coiled. it does not wrap around the octamer smoothly. there are four bends in its helical structure as it winds around the histones. the fifth type of histone is H1, is not a part of the core particle but plays an important role in the nucleosome structure. the precise location of H1 with respect to core particle is still uncertain. H1 sits outside the octamer and binds to the DNA where the DNA joins and leaves the octamer. however the result of recent experiments suggest that the H1 histone sits inside the coils of the nucleosome. H1 helps to lock the DNA into place, acting as a clamp around nucleosome octamer together,the core particle and H1 (linker) are called chromatosome. the next level of chromatin organization. located at regular interval along the DNA molecules and are separated from one another by linker DNA. which varies in size among all cell types most cells have from about 30 to 40 bp of linker DNA. non histone chromosomal proteins may be associated with the linker DNA, and few also appears to bind directly to the core particle. while wrapping around the core histones, the structure of DNA is altered in the middle of nucleosome core particles and exhibit increased number of base pairs per turn.

COMPACTION OF CHROMOSOME

the two events we have discussed so far have shortened the DNA about 50 folds. a third level of compaction involves interaction between 30nm fibre and nuclear matrix. the nuclear matrix is composed of two parts:

nuclear lamina

internal matrix protein

10 nm fibre and associated proteins

NUCLEOSOME JOINS TO FORM A 30NM FIBRE

nucleosome associated with each other to form a more compact structure termed as 30nm fibre. histone H1 plays a role in this compaction. at moderate salt concentrations, H1 is removed. this result is classic beads on string morphology.the 30 nm fibre shortens the total length of DNA  another 7 folds 

its structure has proven difficult to determine

the DNA conformation may be substantially altered when extracted from living cells

two models:

solenoid

zigzag

CHANGES IN CHROMATIN STRUCTURE

eukaryotic  DNA must be tightly packed to fit into the cell nucleus, it must also periodically unwind to undergo replication and transcription. evidence of changing nature of chromatin structure is seen in the puffs of polytene chromosomes and in sensitivity of genes to digestion by DNase I. polytene chromosome are ain't chromosomes found in certain tissues of drosophila and some other organism. these large, unusual chromosomes arise when repeated, rounds of DNA replication takes place without accoumpying cell division, producing thousands of copies of DNA that lie side by side. when polytene chromosomes are stained with dyes, numerous bands are revealed. under certain conditions, the bands may exhibit chromosomal puffs localized swellings of the chromosomes. each puff is a region of chromatin that has relaxed its structure, assuming a more open state. if radioactively labelled uridine is added to  drosophila  larva, radioactivity accumulates in puff, indicating that they are regions of active transcription.appearence of puffs at particular locations on the chromosome can be stimulated by exposure to hormones and other compounds that are known to induce the transcription of genes at loose locations. this correlations between the occurrence of transcription and the relaxation of chromatin at a puff site indicates that chromatin structure undergoes dynamic changes associated with gene activity.

the C 0 t value is the product of C 0 (the initial concentration of DNA), t (time in seconds), and a constant that depends on the concentration of cations in the buffer. Repetitive DNA will renature at low C 0 t values, while complex and unique DNA sequences will renature at high C 0 t values.