Journal
NUCLEIC ACIDS RESEARCH
Volume 36, Issue 2, Pages 559-569Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkm1029
Keywords
-
Categories
Funding
- NCI NIH HHS [P01-CA72765, P01 CA072765, R01-CA101859, R01 CA101859] Funding Source: Medline
- NIDDK NIH HHS [T32-DK0778, T32 DK007780] Funding Source: Medline
Ask authors/readers for more resources
Light-activated antisense oligodeoxynucleotides (asODNs) were developed to control the degradation of target mRNA in living cells by RNase H. A 20-mer asODN previously shown to target c-myb, a hematopoietic transcription factor, was covalently attached via a photocleavable linker (PL) to partially complementary 20-mer sense strands (sODNs). In the 'caged' state, the sODN blocked hybridization of the asODN to c-myb mRNA. Six asODN-PL-sODN conjugates, C1-C6, were synthesized. C5, with twelve complementary bases, gave the largest decrease in melting temperature (T-m) upon UV irradiation (Delta T-m=-29 degrees C). The most thermally stable conjugate, C6 (T-m=84 degrees C), gave the lowest background RNase H activity, with just 8.6% degradation of an RNA 40-mer after 1 h incubation. In biochemical assays with C6, RNA digestion increased 10-fold 10 min after UV irradiation. Finally, phosphor-othioated analogs S-C5 and S-C6 were synthesized to test activity in cultured K562 (human leukemia) cells. No knockdown of c-myb mRNA or protein was observed with intact S-C5 or S-C6, whereas more than half of c-myb mRNA was degraded 24 h after photoactivation. Two-fold photomodulation of c-MYB protein levels was also observed with S-C5. However, no photomodulation of c-MYB protein levels was observed with S-C6, perhaps due to the greater stability of this duplex.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available