Synthetic Biology Parts, Devices and Applications

10.1 An Introduction to Transcript Control 191

The second method of RNase E substrate recognition, often termed “direct
entry,” bypasses the 5′ end [23]. Experiment has demonstrated that mRNAs con-
taining a putative 5′ hairpin to inhibit 5′ RNase E binding is still degraded in an
RNase E-dependent manner [24], and the insertion of putative RNase E sites into
the coding region decreased the stability of RNA with a 5′ hairpin [25].
Additionally, several RNAs have been identified that can be rapidly degraded by
the RNase E catalytic domain even if they are not terminated with a 5′-P (i.e., if

Primary pathways for RNase E-mediated transcript degradation

Unprocessed

mRNA has no

5 ′-P for RNase E

binding

Tr anscription

or ribozyme

cleavage

RNase E

E

E

5 ′-PPP /

5 ′-HO

5 ′-PPP /

5 ′-HO

5 ′ entry Direct entry

(5′-independent)

RppH-mediated No 5′ removal

5 ′-PP removal

RNase E

cleavage

Degradation to

monomers

PNPase Exonucleases

PNP EXO

RNase E

cleavage

5 ′-P

5 ′-P –3′ 5 ′-P

–3′

+ +

–3′

3 ′ 5 ′

degradation

Monomers

+

E

+

–3′

–3′

mRNA

Figure 10.1 Primary routes for RNase E-mediated mRNA degradation. “5′ entry” is initiated
when an mRNA undergoes 5′-PP removal, catalyzed by the pyrophosphohydrolase enzyme
RppH, creating a 5′-P that can be recognized and bound by RNase E (shown by “+” symbol).
“Direct entry” (at right) is 5′ independent entry by RNase E that occurs without recognition
and binding to a 5′-P moiety. Following RNase E binding, an initial cleavage event generates
3 ′-OH- and 5′-P-terminated RNAs that are efficient substrates for 3′ → 5 ′ degradation to
monomers and further rounds of RNase E binding and cleavage.