The generation of a stable, soluble chimeric tripartite motif (TRIM) 5-TRIM21 protein has simplified efforts to study the role of TRIM5 in innate immunity to retroviruses.
TRIM5 is a member of the tripartite motif (TRIM) family of proteins that is involved in innate immunity to retroviruses. In Old World monkeys TRIM5 protects against HIV-1, although this activity is not shared by its human equivalent. Investigations into the mode of action of TRIM5 have been hampered by a lack of ready supplies of pure recombinant protein. Reporting in the Journal of Virology Kar et al. and Langelier et al. report solutions to this problem and present an initial detailed characterization of TRIM5.
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Members of the TRIM family of proteins are involved in a range of cellular functions and typically contain a RING domain, one or more B-box domains and a coiled-coil region. As befits RING-domain containing proteins, several TRIMs have ubiquitin E3 ligase activity. B-box domains are zinc-finger motifs that are only found in TRIM proteins, whereas coiled-coil regions are often involved in protein oligomerization.
Many TRIM proteins also have a SPRY domain at their carboxyl terminus, which, in those TRIM proteins that are involved in innate immunity, determines their specificity for particular retroviruses. The mechanism of this antiviral function might involve ubiquitin ligase activity but TRIM proteins in which the RING domain has been removed still retain antiviral capacity, albeit a capacity that is reduced in strength. The coiled-coil, B-box and SPRY domains are essential for recognizing and combating viruses.
In mammalian cells TRIM5 is quickly degraded and has a tendency to form insoluble aggregates. To avoid these problems, both groups used a version of TRIM5 from rhesus monkeys in which the RING domain was swapped for that of the related human protein, TRIM21. The resulting chimera (TRIM5–21R) was expressed in insect cell cultures. Although aggregation still occurred, TRIM5–21R was purified in both monomeric and dimeric forms, and the dimer seemed to correspond to the authentic oligomeric state of TRIM5 and TRIM5–21R proteins that is expressed in mammalian cells.
Purified TRIM5–21R was capable of auto-ubiquitination, and partnered with several E2 ligases to produce different patterns of mono- and poly-ubiquitination. TRIM5–21R sedimented with model HIV-1 capsids that were assembled from capsid–nucleocapsid fusion proteins, thereby confirming the direct binding of TRIM proteins to viral capsids — an activity that required a 13 amino-acid loop within the SPRY domain, which was implicated in determining the specificity of TRIM5 variants. Using electron microscopy Langelier et al. observed that TRIM5–21R coated 'capsid tubes' and noted that TRIM5–21R increased the number of broken or irregular tubes.
Whether TRIM5 combats viral infection by physically tearing apart the capsids, tagging them for destruction by proteasomes or a combination of these functions is yet to be elucidated, as is the role of oligomerization in these processes. However, this source of pure, recombinant protein should greatly advance our ability to investigate the functions of TRIM5 and all its multifaceted cousins.
Christopher Surridge
References
Kar, A. K., Diaz-Griffero, F., Li, Y., Li, X. & Sodroski, J. Biochemical and biophysical characterization of a chimeric TRIM21–TRIM5 protein. J. Virol.82, 11669–11681 (2008) | Article | PubMed |
Langelier, C. R. et al. Biochemical characterization of a recombinant TRIM5 protein that restricts HIV-1 replication J. Virol.82, 11682–11694 (2008) | Article | PubMed |
Nisole, S., Stoye, J. P. & Saïb, A. TRIM family proteins: retroviral restriction and antiviral defence. Nature Rev. Microbiol.3, 799–808 (2005) | Article | PubMed |
-TRIM21 protein has simplified efforts to study the role of TRIM5
