Polyadenylation signals

Fig. 4. Schematic representation of expression of the rat pyruvate kinase (PK) gene. Exons specific to each isozyme are indicated by marked boxes. The exons common to Mj- and M2-type PK and common to L- and R-type PK are shown by open boxes. CAAT, CAT box TATA, TATA box AATAAA, polyadenylation signal.

PolyA Signal This is the polyadenylation signal for attachment of the polyA tail to generate mature mRNA it is important for transcription termination. 

During polyadenylation the primary transcript is shortened in an endonucleolytic step and appended with ca. 200 A-residues. The endonucleolytic incision requires two signal sequences on the pre-mRNA. A highly conserved AAUAAA sequence 10-30 nucleotides upstream from the hydrolysis site serves as one signal. Another signal in the form of a less well conserved GU- or U-rich element upstream of the hydrolysis site. Both together constitute the polyadenylation signal (Fig. 1.45). Polyadenylation occurs in a multiprotein complex, whose composition is not yet explained in all details. 

Eukaryotic Animal (recombinant) virus vectors Transient or lytic infection Infection in susceptible cells All essential viral genes strong promoter/enhancers polyadenylation signal intron sequences... 

Eukaryotic Mammalian cell-transient expression Transient expression None A higher copy number can be achieved using ori sequence that responds to factors in recipient cells Strong constitutive or inducible promoter polyadenylation signal intron sequences... 

Addition of a poly-A tail Most eukaryotic mRNAs (with several notable exceptions, including those coding for the histones and some interferons) have a chain of 40 to 200 adenine nucleotides attached to the 3 -end (see Rgure 30.17). This poly-A tail is not transcribed from the DNA, but rather is added after transcription by the nuclear enzyme, polyadenylate polymerase. A consensus sequence, called the polyadenylation signal sequence (AAUAAA), found near the 3 -end of the RNA molecule, signals that a poly-A tail is to be added to the mRNA. These tails help stabilize the mRNAs and facilitate their exit from the nucleus. After the mRNA enters the cytosol, the poly-A tail is gradually shortened. 

Most pre-mRNA transcripts are cleaved post-transcriptionally near the 3 end between a polyadenylation signal (5 -AAUAAA-3 ) and 5 -YA-3 (where Y = a pyrimidine). A GU-rich sequence may also be located further downstream. Specific proteins bind to these sequence elements to form a complex. One of the bound proteins, poly(A) polymerase, then adds a poly(A) tail of up to 250 A residues to the new 3 end of the RNA molecule and poly(A) binding protein molecules bind to this. The poly(A) tail protects the 3 end of the final mRNA against nuclease degradation and also increases translational efficiency of the mRNA. Some pre-mRNAs (e.g. histone pre-mRNAs) are cleaved near the 3 end but no poly(A) tail is added. 

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