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1.A.79 The Cholesterol Uptake Protein (ChUP) or Double Stranded RNA Uptake Family

When dsRNA is injected into C. elegans, it spreads to silence gene expression throughout the animal and in its progeny. This phenomenon is termed RNA interference (RNAi) and has been observed in plants and nematodes. SID-1 is a 776 aa residue integral membrane C. elegans protein with a 400 aa extracelular N-terminal domain and a C-terminal domain of 11 putative TMSs that mediates passive dsRNA transport into cells. However, export of RNA silencing from C. elegans tissues does not require SID-1 (Jose et al., 2009). A 9 TMS model with two regions that dip into the membrane from the external side has been proposed (Feinberg and Hunter, 2003). Several distant but probable paralogues of SID-1 are found in C. elegans, and mammals contain SID-1 homologues. It has been shown that a human SID-1 homologue enhances siRNA uptake and gene silencing (Duxbury et al., 2005). A homologue could not be identified encoded within the genome of Drosophila melanogaster or in other organisms.

The human SID-1 homologue FLJ20174 localizes to the cell plasma membrane and enhances uptake of small interfering RNA (siRNA). This results in increased siRNA-mediated gene silencing efficacy. Thus, overexpression enhances siRNA internalization in mammalian cells. The N-terminal extracellular domain of human SID-1 has been characterized (Pratt et al., 2012). It is glycosylated and forms a compact, globular tetramer. It may control access of dsRNA to the transmembrane pore. SID-1 is a dsRNA-selective dsRNA-gated channel (Shih and Hunter, 2011). Both single- and double-stranded dsRNA, such as hairpin RNA and pre-microRNA, can be transported by SID-1.

Survival of C. elegans depends on the dietary absorption of sterols present in the environment. Valdes et al. (2012) provided evidence that Cholesterol Uptake Protein-1 (ChUP-1) (ZK721; tag-130) is involved in dietary cholesterol uptake in C. elegans. Animals lacking ChUP-1 showed hypersensitivity to cholesterol limitation and were unable to uptake cholesterol. A ChUP-1-GFP fusion protein colocalized with cholesterol-rich vesicles, endosomes and lysosomes as well as the plasma membrane. A direct interaction was found between the cholesterol analog DHE and the transmembrane 'cholesterol recognition/interaction amino acid consensus' (CRAC) motif present in C. elegans ChUP-1. In-silico analysis identified two mammalian homologues of ChUP-1. CRAC motifs are conserved in mammalian ChUP-1 homologues (Valdes et al., 2012). 

Single-stranded oligonucleotides (ssOligos) are efficiently taken up by living cells without the use of transfection reagents. This phenomenon, called 'gymnosis', enables the sequence-specific silencing of target genes. Several antisense ssOligos are used for the treatment of human diseases. Systemic RNA interference deficient-1 (SID-1) transmembrane family 2 (SIDT2), a mammalian ortholog of the Caenorhabditis elegans double-stranded RNA channel SID-1, mediates gymnosis. Takahashi et al. 2017 showed that the uptake of naked ssOligos into cells is downregulated by knockdown of SIDT2, and it inhibited the effect of antisense RNA mediated by gymnosis. Overexpression of SIDT2 enhanced the uptake of naked ssOligos into cells, while a single amino acid mutation in SIDT2 abolished this effect. Thus, SIDT2 mediates extra- and intracellular RNA transport.


Transport reactions believed to be catalyzed by SID-1 and ChUP1 are:


dsRNA (out) ⇌ dsRNA (in)

Cholesterol (out)  ⇌  Cholesterol (in)

A transport reaction believed to be catalyzed by SIDT2 of mammals is:

ssRNAout (oligonucleoties) ⇌ ssRNAin (oligonucleotides)

References associated with 1.A.79 family:

Aizawa, S., V.R. Contu, Y. Fujiwara, K. Hase, H. Kikuchi, C. Kabuta, K. Wada, and T. Kabuta. (2016). Lysosomal membrane protein SIDT2 mediates the direct uptake of DNA by lysosomes. Autophagy 0. [Epub: Ahead of Print] 27846365
Aizawa, S., Y. Fujiwara, V.R. Contu, K. Hase, M. Takahashi, H. Kikuchi, C. Kabuta, K. Wada, and T. Kabuta. (2016). Lysosomal putative RNA transporter SIDT2 mediates direct uptake of RNA by lysosomes. Autophagy 12: 565-578. 27046251
Cappelle, K., C.F. de Oliveira, B. Van Eynde, O. Christiaens, and G. Smagghe. (2016). The involvement of clathrin-mediated endocytosis and two Sid-1-like transmembrane proteins in double-stranded RNA uptake in the Colorado potato beetle midgut. Insect Mol Biol. [Epub: Ahead of Print] 26959524
Contu, V.R., K. Hase, H. Kozuka-Hata, M. Oyama, Y. Fujiwara, C. Kabuta, M. Takahashi, F. Hakuno, S.I. Takahashi, K. Wada, and T. Kabuta. (2017). Lysosomal targeting of SIDT2 via multiple YXXΦ motifs is required for SIDT2 function in the process of RNautophagy. J Cell Sci. [Epub: Ahead of Print] 28724756
Dickson, E.J., J.B. Jensen, O. Vivas, M. Kruse, A.E. Traynor-Kaplan, and B. Hille. (2016). Dynamic formation of ER-PM junctions presents a lipid phosphatase to regulate phosphoinositides. J. Cell Biol. [Epub: Ahead of Print] 27044890
Duxbury, M.S., S.W. Ashley, and E.E. Whang. (2005). RNA interference: a mammalian SID-1 homologue enhances siRNA uptake and gene silencing efficacy in human cells. Biochem. Biophys. Res. Commun. 331: 459-463. 15850781
Feinberg, E.H. and C.P. Hunter. (2003). Transport of dsRNA into cells by the transmembrane protein SID-1. Science 301: 1545-1547. 12970568
Herdman, C. and T. Moss. (2016). Extended-Synaptotagmins (E-Syts); the extended story. Pharmacol Res 107: 48-56. [Epub: Ahead of Print] 26926095
Jose, A.M., J.J. Smith, and C.P. Hunter. (2009). Export of RNA silencing from C. elegans tissues does not require the RNA channel SID-1. Proc. Natl. Acad. Sci. USA 106: 2283-2288. 19168628
Jose, A.M., Y.A. Kim, S. Leal-Ekman, and C.P. Hunter. (2012). Conserved tyrosine kinase promotes the import of silencing RNA into Caenorhabditis elegans cells. Proc. Natl. Acad. Sci. USA 109: 14520-14525. 22912399
McEwan, D.L., A.S. Weisman, and C.P. Hunter. (2012). Uptake of extracellular double-stranded RNA by SID-2. Mol. Cell 47: 746-754. 22902558
Pratt, A.J., R.P. Rambo, P.W. Lau, and I.J. MacRae. (2012). Preparation and characterization of the extracellular domain of human Sid-1. PLoS One 7: e33607. 22509261
Shih, J.D. and C.P. Hunter. (2011). SID-1 is a dsRNA-selective dsRNA-gated channel. RNA 17: 1057-1065. 21474576
Takahashi, M., V.R. Contu, C. Kabuta, K. Hase, Y. Fujiwara, K. Wada, and T. Kabuta. (2017). SIDT2 mediates gymnosis, the uptake of naked single-stranded oligonucleotides into living cells. RNA Biol 0. [Epub: Ahead of Print] 28277980
Valdes, V.J., A. Athie, L.S. Salinas, R.E. Navarro, and L. Vaca. (2012). CUP-1 Is a Novel Protein Involved in Dietary Cholesterol Uptake in Caenorhabditis elegans. PLoS One 7: e33962. 22479487
Xu J., Yoshimura K., Mon H., Li Z., Zhu L., Iiyama K., Kusakabe T. and Lee JM. (2014). Establishment of Caenorhabditis elegans SID-1-dependent DNA delivery system in cultured silkworm cells. Mol Biotechnol. 56(3):193-8. 23979877
Xu, W. and Z. Han. (2008). Cloning and phylogenetic analysis of sid-1-like genes from aphids. J Insect Sci 8: 1-6. 20302524
Yu, H., Y. Liu, D.R. Gulbranson, A. Paine, S.S. Rathore, and J. Shen. (2016). Extended synaptotagmins are Ca2+-dependent lipid transfer proteins at membrane contact sites. Proc. Natl. Acad. Sci. USA. [Epub: Ahead of Print] 27044075