In this article we will discuss about:- 1. Meaning of Crossing Over 2. Frequency of Crossing Over 3. Factors 4. Molecular Mechanism.
Meaning of Crossing Over:
Crossing over may be defined as an interchange of corresponding chromosomal parts between members of a homologous chromosome resulting in a recombination of genes. One, two or more fragments may be interchanged during crossing over.
The phenomenon crossing over generally occurs during meiosis, particularly between leptotene to zygotene stages of Prophase 1 and generally takes place at the four strand stage.
Frequency of Crossing Over:
How often crossing over occurs between two genes determines the frequency of crossing over or the percentage of crossing over for those two genes. Normally the percentage of crossing over is directly proportional to the distance between the two genes i.e. percentage is high if the distance is great and vice versa.
Therefore, the percentage of crossing over between two genes indicates the distance between two genes. Haldane termed the unit of crossing over as “morgan” and thus if 2.5% crossing over occurs between two genes then they are said to be 2.5 morgan unit apart.
Factors Controlling the Frequency of Crossing Over:
i) High and low temperature increase the frequency of crossing over.
ii) X-rays also increase the frequency of crossing over.
iii) Older the age, greater the frequency of crossing over.
iv) Gene mutation also affects the frequency of crossing over.
v) Irradiation by radium also increases the frequency of crossing over.
vi) Sex of the individual also has an influence in the frequency of crossing over.
vii) Heterochromatic region is less affected than euchromatic region.
viii) Inversion and other chromosomal abberration also suppress the crossing over.
In this regard it should be remembered that crossing over is not an accidental process but an essential cellular process catalysed by enzymes that cell encode and regulate for the purpose.
Molecular Mechanism of Crossing Over:
Until recently there was not even an superficial understanding of the molecular basis of crossing over. During 1930, from the cytological observations, it was hypothesized that during meiosis the paired coiled chromosomes were sometimes physically broken at the chromatin level as a result of tension created by their contraction.
After that the broken ends reunites in crossways phenomenon creating two reciprocally recombinant chromatids. According to this model, recombination occurs after chromosome duplication is complete. But this hypothesis fell into disfavour during 1955 when geneticist found that crossing over occurs within the gene.
As a result of this dilemma an alternative hypothesis was also proposed and according to this hypothesis during the time of replication of paired chromosomes the new DNA strands being formed along the paternal chromosome switches to the maternal one and thereafter copying. This hypothesis was known as “copy choice hypothesis”.
A fundamental distinction between the two hypotheses lies in their prediction of physical origin of recombinant chromosomes. But recently different experimental evidences revealed that recombination can occur between non-replicating DNA molecules, therefore breakage and reunion of – intact double helix may be the primary mechanism of crossing over.
Homologous crossing over is guided by base-pairing:
Crossing over occurs between two regions of DNA containing identical or nearly identical sequences.
Crossing over is initiated from breaks or gaps in DNA:
Breaks in the DNA greatly stimulates the crossing over. Direct proof of this statement is that a plasmid bearing a yeast DNA segment can be made to recombine with the yeast chromosome up to a thousand times more efficiently by cutting the yeast segment with a restriction enzyme before the plasmid is introduced into the cell. Similarly, cellular enzymes actively induce recombination in meiosis by making occasional breaks along a pair of synapsed chromosome.
Rec-A protein is the initiator of recombination:
Rec-A is an enzyme that carries out the fundamental reaction in recombination by joining two DNA molecules. The unique properties of Rec-A protein is that it recognises single stranded DNA and anneals it to a complementary sequence in a homologous duplex. Rec-A protein first binds single stranded DNA in a fixed ratio of about one Rec-A protein per five nucleotide and thus forming a DNA- protein filament.
If ATP is present then Rec-A protein in the filament may unwind successively different parts of the duplex attempting to anneal the bound single strand to unwound regions until a complementary sequence is found. The energy of ATP drives the pairing reaction. After a new hybrid DNA formation, Rec-A protein is displaced.
Crossing over involves the cross strand intermediate:
During the time of crossing over an intermediate is formed in which two DNA duplex join in recombination in such a fashion that a cross strand junction is formed which is known as “holliday-structure”—previously known as “chiasmata”.
Formation of Heteroduplex:
Heteroduplex are regions on recombinant DNA molecules where the two strands are not exactly complementary. The frequent generation of heteroduplex segments during crossing over provides direct experimental support for the hypothesis that the fundamental recombination even involves pairing between regions of single stranded DNA. Heteroduplex lifetime is very short because, when the recombinant DNA molecules duplicate, the alternative alleles segregate out.
Rec-BC enzyme promotes recombination by unwinding DNA and nicking it at specific sites known as ‘Chi’:
This rec-BC enzyme has a molecular weight of about 3, 00,000 dalton. This enzyme has both DNA unwinding and nuclease activity. An important feature of Rec-BC enzyme is that it initiates unwinding only on the DNA containing a free duplex end. This Rec-BC enzyme uses energy to travel along the duplex for unwinding. One unique property of Rec-BC enzyme is that it unwinds DNA faster than it rewinds.
Therefore unwound segments become progressively longer as the enzyme moves down the DNA. For this reason the unwound structures form two single stranded loops known as “rabbit ears”. Genetic experiments by Kobayashi et al (1984) and Ponticelli et al (1985) showed that Rec-BC enzyme promotes recombination most frequently in DNA containing a site known as ‘Chi’ which’ has the nucleotide sequence 5′ GCTGGTGG 3′.
When ‘Chi’ is present in DNA being unwound a specific nuclease activity of Rec-BC enzyme cuts the exposed single strand near ‘Chi’ thus preventing it being rewound as the enzyme progresses. The consequence is to leave behind a free single stranded tail and a gap, both of which are sites where Rec-A protein can bind to initiate DNA strand exchange.
Recombination is not always reciprocal at the site of crossing over:
Generally, crossing over results in a reciprocal recombination but exceptions were discovered where non-reciprocal recombination occurs. This non-reciprocal recombination during crossing over is also known as gene conversion.
Mechanism of Calculating the Frequency of Crossing Over:
By using the following mathematical notation we can calculate the frequency of recombination:
Frequency of recombination (R) = Sum of two recombinant types / Sum of all types x 100
We can also calculate the co-efficient of coincidence in case of double or triple or more crossing over by using the following principle:
Co-efficient of coincidence (S) = Recombination 1,2………………………../(Recombination 1) x (Recombination 2) x……………………
After calculation if:
S = 1, it indicates one has no influence on the other
S = fractional number, double crossing over is less than predicted
S = greater than 1, double crossing over exceeds expectations.
Crossing over creates a genetic diversity by producing different combinations of particular genetic alleles and also allows repair of DNA damaged by radiation or other chemicals through reconstruction of intact DNA from segments.