Additional information

1. Telomeres and telomerase: Telomeres are DNA–protein structures located at both ends of each chromosome, consisting of tandem repeats of TTAGGG. A small portion of telomeric DNA is lost with each cell division. When telomere length reaches a critical limit, the cell undergoes senescence and/or apoptosis. Telomere length is maintained by the enzyme telomerase which adds “TTAGGG” repeats to chromosome ends. Telomeres are essential for the telophase stage of mitosis. When telomeres becomes too short, mitosis and cell division cannot occur.
2. Rb and p53 tumor suppressor proteins*

• Rb or retinoblastoma protein: Rb prevents the cell from entering into S phase from G1 phase. Phosphorylated Rb is inactive. Rb normally binds to transcription factor E2F and inhibits it. In the presence of growth factors, cyclin D-CDK4/6 and cyclin E-CDK2 phosphorylate and inactivate Rb. As a result, E2F is released and it binds DNA to and increases transcription of S phase genes like DNA polymerase. Rb gene is mutated in hereditary retinoblastoma.
• p53 protein: It prevents cell transition from G1 to S. p53 is activated in the presence of DNA damage. p53, as a transcription activator, turns on the expression of genes like P21 and P27, that inhibit the cell cycle. It induces apoptosis in cells with severe DNA damage. In normal cells, Mdm 2 keeps the levels of p53 low. DNA damage activates a set of kinases that phosphorylate Mdm2, causing it to dissociate from p53. Mutations in p53 lead to unregulated cell division and cancers.

* HPV 16 and 18 viral proteins E6 and E7 bind to and inhibit p53 and Rb respectively, leading to malignancies like cervical cancers.

3. ATM and ATR

ATM and ATR are tumor suppressor proteins that inactivate cdc25C phosphatase and prevent the cell from proceeding to M phase. ATM mediates the cell response to double stranded DNA breaks induced by ionizing radiation. Mutations in ATM gene are seen in ataxia telangiectasia. ATR mediates the cell response to UV induced DNA damage and double stranded DNA breaks.

4. Extracellular vesicles (EVs) are small-membrane vesicles secreted by most cell types with the role to provide intercellular communication both locally and systemically. Cells may use EVs to transfer nucleic acids, proteins and lipids. EVs play a role in tumor progression and metastases.

   The transfer of their content between cells, which includes confers the means for these interactions and induces significant cellular behaviour changes in the receiving cell. EVs are implicated in the regulation of numerous physiological and pathological processes, including development and neurological and cardiovascular diseases. Importantly, it has been shown that EV signalling is essential in almost all the steps necessary for the progress of carcinomas, from primary tumours to metastasis. EVs help tumors in angiogenesis, invasion, metastasis and conversion of local fibroblasts into cancer associated fibroblasts. In Ca breast metastasis to the brain, EVs are taken up by the BBB by transcytosis.

5. DNA is read from upstream to downstream i.e. from 3’ to 5’ ends.

6. t RNA (transfer RNA): It transfers amino acids to the growing peptide chain. It has a folded structure with three hairpin loops that look like a three-leaf clover. It contains an anticodon that is complementary to the mRNA codon. The 3’ end of tRNA is called the acceptor arm and binds to the amino acid. The enzyme aminoacyl-tRNA synthetase adds an amino acid to its specific tRNA which “charges” the tRNA. This enzyme also has proofreading activity making sure that the correct amino acid binds to its specific tRNA.

7. How to write complementary DNA sequences? Because of the nature of complementary base pairing, if you know the sequence of one strand of DNA, you can predict the sequence of the strand that will pair with, or “complement” it. Remember, when writing complementary DNA sequences, you need to write the sequence in the 5’ to 3’ direction. This usually involves reversing the sequence.

   e.g. what will be the complementary DNA sequence to ATTGCGA ? As per protocol, we can assume that it is written in 5’ to 3’, which is 5’ ATTGCGA 3’. Strictly looking at complementary base pairing the sequence will be 3’ TAACGCT 5’. Now, reverse it to write in 5’ to 3’ protocol, hence the answer will be 5’ TCGCAAT 3’ also written simply as TCGCAAT. When writing an RNA sequence replace T with U (for uracil).

8. Protooncogenes and tumor suppressor genes

9. Common tumor markers