There are two major steps to start translation

1. Activation of amino acids,

2. Assembly of ribosomes.

TLS Online TPP Program

#Id: 6503

#Unit 3. Fundamental Processes

These two mechanisms, (1) binding of the capping enzyme to RNA polymerase II specifically through its unique CTD phosphorylated on serine 5 of the heptapeptide repeat during transcription initiation by TFIIH, and (2) activation of the capping enzyme by the serine 5–phosphorylated CTD, result in specific capping of RNAs transcribed by RNA polymerase II.

TLS Online TPP Program

#Id: 6504

#Unit 3. Fundamental Processes

Once bound to Pol II, Ptefb phosphorylates the serine residue at position 2 of the CTD repeats. That phosphorylation event correlates with elongation. In addition, P-TEFb phosphorylates and thereby activates another protein, called SPT5, itself an elongation factor. Finally, TAT-SF1, yet another elongation factor, is recruited by P-TEFb. Thus, P-TEFb stimulates elongation in three separate ways.

TLS Online TPP Program

#Id: 6755

#Unit 3. Fundamental Processes

Most mRNAs decay stochastically (like the decay of radioactive isotopes), and as a result mRNA stability is usually expressed as a half- life (t½).

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#Id: 6756

#Unit 3. Fundamental Processes

Method for determining mRNA half-lives.

RNA polymerase II transcription is shut down, either by a drug or a temperature shift in strains with a temperature sensitive mutation in a Pol II gene. The levels of specific mRNAs are determined by northern blot or RT-PCR at various times following shutdown.

TLS Online TPP Program

#Id: 6757

#Unit 3. Fundamental Processes

In E. coli the typical mRNA half-life is about 3 minutes, but half-lives of individual mRNAs may be as short as 20 seconds or as long as 90 minutes.  In budding yeast, mRNA half-lives range from 3 to 100 minutes, whereas in metazoans, half-lives range from minutes to hours, and in rare cases, even days.

TLS Online TPP Program

#Id: 6758

#Unit 3. Fundamental Processes

Degradation of bacterial mRNAs is initiated by removal of a pyrophosphate from the 5’ terminus. The monophosphorylated form stimulates the catalytic activity of an endonuclease RNase E