Mechansism of cellular uptake of modified oligodeoxynucleotides containing methylphosphonate linkages

Yoko Shoji, Saghir Akhtar, Ammasi Periasamy, Brian Herman, R. L. Juliano

Research output: Contribution to journalArticlepeer-review

166 Scopus citations

Abstract

The cellular uptake and intracellular distribution of methylphosphonate oligonucleotides (15 mers) has been examined using both 32P labeled and luorescent labeled oligonucleotides. The cellular uptake process for methylphosphonate oligonucleotides is highly temperature dependent, with a major increase in uptake occuring between 15 and 20°C. Most of the label which becomes cell associated at 37°C cannot be removed by acid washing or trypsinization and thus seems to be within the cell. Visualization of rhodamine labeled methylphosphonate oligonucleotides using digital imaging fluorescence microscopy reveals a vesicular subcellular distribution suggestive of an endosomal localization. There was extensive co-localization of rhodamine labeled methylphosphonate oligonucleotides with fluorescein-dextran, an ndosomal/lysosomal arker substance. The apparent endocytotic uptake of labeled methylphosphonate oligonucleotides could not be blocked by competition with unlabeled methylphosphonate or phosphodiester oligonuclotides, nor by ATP. This contrasts with the situation for radiolabeled phosphodiester oligonucleotides whose uptake can be completely blocked with unlabeled competitor. Uptake of phosphodiester oligonucleotides, but not of methylphosphonate oligonucleotides, could be locked by acidification of the cytosol. These observations suggest that the pathway of cellular uptake of methylphosphonate oligonucleotides involves fluid phase or adsorbtive endocytosis, and is distinct from the uptake pathway for phosphodiester oligonucleotides.

Original languageEnglish (US)
Pages (from-to)5543-5550
Number of pages8
JournalNucleic acids research
Volume19
Issue number20
DOIs
StatePublished - Oct 25 1991

Bibliographical note

Funding Information:
This work was supported by NIH grant

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