These papers use Morpholinos targeted to unusual sites on RNA. Some target non-coding RNA at splice junctions, protein binding sites or base-pairing moieties.
Hu Z, Chen G, Zhao Y, Gao H, Li L, Yin Y, Jiang J, Wang L, Mang Y, Gao Y, Zhang S, Ran J, Li L. Exosome-derived circCCAR1 promotes CD8 + T-cell dysfunction and anti-PD1 resistance in hepatocellular carcinoma. Mol Cancer. 2023 Mar 18;22(1):55. doi:10.1186/s12943-023-01759-1.
Target is a methylation site (m6A) on a cccRNA
Liu Y, Yang Y, Lin Y, Wei B, Hu X, Xu L, Zhang W, Lu J. N6 -methyladenosine-modified circRNA RERE modulates osteoarthritis by regulating β-catenin ubiquitination and degradation. Cell Prolif. 2022 Jun 22:e13297. doi: 10.1111/cpr.13297. Online ahead of print.
"Thus, morpholino oligos (MOs) targeting two HRSP12-binding motifs (GGUUC) in circRERE (MOs-circRERE) were designed by browsing its sequence... Transfection with MOs-circRERE partially reversed the decreased expression of circRERE in HCs stimulated with IL-1β for 6 h, but not 12 or 36-h" (human primary chondrocytes, delivery by Endo-Porter). "However, mice IA-injected with vivo-morpholino targeting GGUUC motif in circRere (vivo-MO-circRere) did not exhibit reductions in the examined manifestations of DMM-induced OA 5-weeks after DMM"
Liu Z, Wang T, She Y, Wu K, Gu S, Li L, Dong C, Chen C, Zhou Y. N(6)-methyladenosine-modified circIGF2BP3 inhibits CD8(+) T-cell responses to facilitate tumor immune evasion by promoting the deubiquitination of PD-L1 in non-small cell lung cancer. Mol Cancer. 2021 Aug 20;20(1):105. doi: 10.1186/s12943-021-01398-4.
"By using sequence-specific morpholino antisense oligos (MAOs) targeting the m6A site of circIGF2BP3, we found that NSCLC cells treated with MAO targeting circIGF2BP3 (MAO-circIGF2BP3) had drastically suppressed circIGF2BP3 expression and decreased m6A levels of circIGF2BP3 compared with cells treated with control MAO (MAO-NC)"
Saha K, Fernandez MM, Biswas T, Joseph S, Ghosh G. Discovery of a pre-mRNA structural scaffold as a contributor to the mammalian splicing code. bioRxiv. 2021;[preprint] doi:10.1101/292458
"morpholino oligonucleotide (AMO) complementary to the 5′ end of U1 snRNA"
Huang C, Zhu B, Leng D, Ge W, Zhang XD. Long noncoding RNAs implicated in embryonic development in Ybx1 knockout zebrafish. FEBS Open Bio. 2021 Apr;11(4):1259-1276. doi: 10.1002/2211-5463.13057. Epub 2021 Feb 26.
Xu J, Ji L, Liang Y, Wan Z, Zheng W, Song X, Gorshkov K, Sun Q, Lin H, Zheng X, Chen J, Jin RA, Liang X, Cai X. CircRNA-SORE mediates sorafenib resistance in hepatocellular carcinoma by stabilizing YBX1. Signal Transduct Target Ther. 2020 Dec 26;5(1):298. doi: 10.1038/s41392-020-00375-5.
Blocking a protein binding site on a circRNA
Xu J, Wan Z, Tang M, Lin Z, Jiang S, Ji L, Gorshkov K, Mao Q, Xia S, Cen D, Zheng J, Liang X, Cai X. N6-methyladenosine-modified CircRNA-SORE sustains sorafenib resistance in hepatocellular carcinoma by regulating β-catenin signaling. Mol Cancer. 2020:19(1):163. doi:10.1186/s12943-020-01281-8
Blocking an adenosine methylation site (N6) in a circular RNA with a Morpholino
Liu X, Wang X, Li J, Hu S, Deng Y, Yin H, Bao X, Zhang QC, Wang G, Wang B, Shi Q, Shan G. Identification of mecciRNAs and their roles in the mitochondrial entry of proteins. Sci China Life Sci. 2020;[Epub ahead of print] doi:10.1007/s11427-020-1631-9
Morpholinos targeting circular RNAs
Gao T, Li J, Li N, Gao Y, Yu L, Zhuang S, Zhao Y, Dong X. lncrps25 play an essential role in motor neuron development through controlling the expression of olig2 in zebrafish. J Cell Phys. 2019;[Epub ahead of print] doi:10.1002/jcp.29237
Targeting an lncRNA
Herrera-Úbeda C, Marín-Barba M, Navas-Pérez E, Gravemeyer J, Albuixech-Crespo BA, Wheeler GN, Garcia-Fernàndez J. Microsyntenic Clusters Reveal Conservation of lncRNAs in Chordates Despite Absence of Sequence Conservation. Biology. 2019;8(3):61. doi:10.3390/biology8030061
Targeting splicing of an lncRNA
Wu M, Zhang S, Chen X, Xu H, Li X. Expression and function of lncRNA MALAT-1 in the embryonic development of zebrafish. Gene. 2018 Sep 21. pii: S0378-1119(18)30996-X. doi: 10.1016/j.gene.2018.09.037. [Epub ahead of print]
Targeting a noncoding RNA, MALAT-1.
Christov CP, Dingwell KS, Skehel M, Wilkes HS, Sale JE, Smith JC, Krude T. A NuRD Complex from Xenopus laevis Eggs Is Essential for DNA Replication during Early Embryogenesis. Cell Rep. 2018;22(9):2265-2278. doi:10.1016/j.celrep.2018.02.015
Targeting a noncoding RNA, Y3.
Chowdhury TA, Koceja C, Eisa-Beygi S, Kleinstiver BP, Kumar SN, Lin CW, Li K, Prabhudesai S, Joung JK, Ramchandran R. Temporal and Spatial Post-Transcriptional Regulation of Zebrafish Tie1 mRNA by Long Noncoding RNA During Brain Vascular Assembly. Arterioscler Thromb Vasc Biol. 2018 May 3. pii: ATVBAHA.118.310848. doi: 10.1161/ATVBAHA.118.310848. [Epub ahead of print].
Lu Y, Hu Z, Mangala LS, Stine ZE, Hu X, Jiang D, Xiang Y, Zhang Y, Pradeep S, Rodriguez-Aguayo C, Lopez-Berestein G, Demarzo A, Sood AK, Zhang L, Dang CV. MYC targeted long non-coding RNA DANCR promotes cancer in part by reducing p21 levels. Cancer Res. 2017;[Epub ahead of print] doi:10.1158/0008-5472.
Targeting lncRNA with Vivo-Morpholinos targeting splicing in P493-6 cell culture
Garcia GR, Goodale BC, Wiley MW, La Du JK, Hendrix DA, Tanguay RL. In vivo characterization of an AHR-dependent long non-coding RNA required for proper Sox9b expression. Mol Pharmacol. 2017 Apr 6. pii: mol.117.108233. doi: 10.1124/mol.117.108233. [Epub ahead of print]
MO modified splicing of a ncRNA.
Crossley MP, Krude T. Targeting Functional Noncoding RNAs. Methods Mol Biol. 2017;1565:151-160. doi: 10.1007/978-1-4939-6817-6_13.
Costales MG, Rzuczek SG, Disney MD. Comparison of small molecules and oligonucleotides that target a toxic, non-coding RNA. Bioorg Med Chem Lett. 2016 Apr 11. pii: S0960-894X(16)30386-9. doi: 10.1016/j.bmcl.2016.04.025. [Epub ahead of print]
Kurian L, Aguirre A, Sancho-Martinez I, Benner C, Hishida T, Nguyen TB, Reddy P, Nivet E, Krause MN, Nelles DA, Esteban CR, Campistol JM, Yeo GW, Belmonte JCI. Identification of Novel Long Noncoding RNAs Underlying Vertebrate Cardiovascular Development. Circulation. 2015;131:1278-1290. doi:10.1161/CIRCULATIONAHA.114.013303
Lee N, Moss WN, Yario TA, Steitz JA. EBV noncoding RNA binds nascent RNA to drive host PAX5 to viral DNA. Cell. 2015 Feb 12;160(4):607-18. doi: 10.1016/j.cell.2015.01.015. Epub 2015 Feb 5.
A Morpholino prevents a ncRNA from interacting with a transcript.
Kowalski MP, Baylis HA, Krude T. Non-coding stem-bulge RNAs are required for cell proliferation and embryonic development in C. elegans. J Cell Sci. 2015 Jun 1;128(11):2118-29. doi: 10.1242/jcs.166744. Epub 2015 Apr 23.
Li IC, Chen YC, Wang YY, Tzeng BW, Ou CW, Lau YY, Wu KM, Chan TM, Lin WH, Hwang SP, Chow WY. Zebrafish Adar2 Edits the Q/R Site of AMPA Receptor Subunit gria2α Transcript to Ensure Normal Development of Nervous System and Cranial Neural Crest Cells. PLoS One. 2014 May 12;9(5):e97133. doi: 10.1371/journal.pone.0097133. eCollection 2014.
A Morpholino oligo "designed to complement the sequence of exon complementary sequence (ECS) within the intron downstream to the Q/R editing site of gria2α was used to block the Q/R editing of gria2α."
Wheeler T, Leger AJ, Pandey SK, MacLeod AR, Nakamori M, Cheng SH, Wentworth BM, Bennett CF, Thornton CA. Targeting nuclear RNA for in vivo correction of myotonic dystrophy. Nature. 2012;488:111-5 doi:10.1038/nature11362.
Target: RNA repeat element
van Dijk M, Thulluru HK, Mulders J, Michel OJ, Poutsma A, Windhorst S, Kleiverda G, Sie D, Lachmeijer AMA, Oudejans CBM. HELLP babies link a novel lincRNA to the trophoblast cell cycle. J Clin Invest. 2012;[Epub ahead of print] doi:10.1172/JCI65171.
Morpholinos targeting mutant sites causing HELLP; the "HELLP locus is in an intergenic region on 12q23.2 between PMCH and IGF1" with mutations on "a novel long intergenic noncoding RNA (lincRNA) transcript of 205,012 bases with (peri)nuclear expression in the extravillous trophoblast".
Ulitsky I, Shkumatava A, Jan CH, Sive H, Bartel DP. Conserved function of lincRNAs in vertebrate embryonic development despite rapid sequence evolution. Cell. 2011 Dec 23;147(7):1537-50.
"One strategy was to inject MOs designed to target lincRNA splice sites in an attempt to disrupt maturation. The other strategy was to inject MOs designed to target highly conserved sites presumed to be important for interactions with other cellular factors." "Embryos injected with either splice- or conserved-site MOs exhibited similar developmental defects"
Chu C, Qu K, Zhong FL, Artandi SE, Chang HY. Genomic Maps of Long Noncoding RNA Occupancy Reveal Principles of RNA-Chromatin Interactions. Mol Cell. 2011 Nov 18;44(4):667-78. doi: 10.1016/j.molcel.2011.08.027. Epub 2011 Sep 29.
"We initially attempted to capture an lncRNA with [three] morpholino probes..." "...with the three-probe approach we could retrieve at most ∼10% of HOTAIR RNA." "Thus, we designed 48 complementary DNA oligonucleotides that were 20-mer each and tiled the entire length of HOTAIR." "With the tiling probes, we could pull down almost all HOTAIR RNA from chromatin..."
Higa-Nakamine S, Suzuki T, Uechi T, Chakraborty A, Nakajima Y, Nakamura M, Hirano N, Suzuki T, Naoya Kenmochi N. Loss of ribosomal RNA modification causes developmental defects in zebrafish. Nucl. Acids Res. 2011. [Epub ahead of print September 8] doi:10.1093/nar/gkr700.
"sqRT–PCR revealed that the U26 snoRNA precursor accumulated in the U26MOsp morphants, which indicates that the MO disrupted host gene splicing. Northern blot analysis showed decreased mature U26 snoRNA expression, but unaltered mature U22 and U27 expression in these morphants, which indicates that the U26MOsp specifically inhibited U26 snoRNA synthesis ... . Similarly, zebrafish embryos injected with U44MOsp showed an accumulation of the U44 precursor transcript and a decrease in mature U44 snoRNA expression ... ."
Collart C, Christov CP, Smith JC, Krude T. The mid-blastula transition defines the onset of Y RNA-dependent DNA replication in Xenopus laevis. Mol Cell Biol. 2011 Jul 25. [Epub ahead of print].
"Injected embryos developed normally until MBT and then showed a delay in development and subsequently died. Embryos injected with xY3MO died soon after MBT at stage 9. Embryos injected with xY4MO and xY5MO showed delayed gastrulation and failed to develop a neural fold. They did not survive beyond the time at which control embryos had reached early neurula and tail bud stages, respectively (Fig. 2A). Injection with an MO against xYα RNA, which is expressed at much lower levels, did not detectably affect early development ... . "