Rat CRM1 Is Responsible for the Poor Activity of Human T-Cell Leukemia Virus Type 1 Rex Protein in Rat Cells

Abstract

Rat models of human T-cell leukemia virus type 1 (HTLV-1)-related diseases such as adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis have been reported. However, these models do not completely reproduce human diseases partly because HTLV-1 replicates poorly in rats. We investigated here the possible reason for this. We found that the activity of Rex in rat cells is quite low compared to that in human cells. As Rex function depends largely on the CRM1 protein, whose human type (human CRM1 [hCRM1]) directly binds to Rex and exports it from the nucleus to the cytoplasm, we assessed whether rat CRM1 (rCRM1) could act as well as hCRM1 as a cofactor for Rex activity. We first cloned a cDNA encoding rCRM1 and found that both rCRM1 and hCRM1 could bind to and export Rex protein to the cytoplasm with similar efficiencies. However, unlike hCRM1, rCRM1 could hardly support Rex function because of its poor ability in inducing the Rex-Rex interaction required for RNA export into the cytoplasm. These observations suggest that the poor ability of rCRM1 to act as a cofactor for Rex function may be responsible for the poor replication of HTLV-1 in rats.

CRM1, an RNA transporter, is a major species-specific restriction factor of human T cell leukemia virus type 1 (HTLV-1) in rat cells

“…The cells (1x10 5 ) were transfected with 0.4 µg of pDM128RxRE [33], 0.05µg of pSRαRex, and 0.1µg of pCDMβ-Gal, Twenty-four hours after transfection the cells were lyzed and the amount of CAT was quantified using a CAT ELISA kit (Roche). The β−galactosidase (β−gal) activity was measured by standard colorimetric methods, to normalize the transfection efficiency, The Rex activity was represented by ratio of CAT/ β−gal for each sample as described previously [33].…”

“…The β−galactosidase (β−gal) activity was measured by standard colorimetric methods, to normalize the transfection efficiency, The Rex activity was represented by ratio of CAT/ β−gal for each sample as described previously [33].…”

“…Immunoblot analysis was performed for detection of CRM1 using affinity purified chicken anti-hCRM1 and rabbit anti-rCRM1 antibodies [33]. To detect HTLV-1 proteins, we used rabbit anti Rex antisera, mouse anti-Tax MAb Lt-4, and mouse anti-p24 Gag MAb NOR-1 [42].…”

“…Rat CRM1 induces minimal amount of Rex multimerization on cognate RNA, although it efficiently exports Rex protein to the cytoplasm. This may cause the defect in viral RNA transport [33]. Two residues (amino acids 411 and 414) in the central region of human CRM1 are crucial for multimerization [34].…”

“…These results suggest that a transgenic (Tg) rat, which expresses human CRM1, may be a model animal to support replication of HTLV-1.Prior to constructing the Tg rat, we examined the effects of hCRM1, expressed at physiological levels in rat cells, because the above results were obtained by overexpression of human and rat CRM1 and toxic effects from overexpressed CRM1 and a dominant-negative influence of rCRM1 over hCRM1 have been reported [33,35]. Moreover, an understanding of the entire viral life cycle is needed.…”