Functional Diversity of Mx Proteins: Variations on a Theme of Host Resistance to Infection

In vertebrates, host defense against pathogens is mediated by two general systems: innate and acquired immunity. Innate immunity constitutes the first line of defense, providing a rapid response by the expression of germ-line encoded proteins that preexist or are induced within hours of infection. Adaptive immunity is a slower, yet highly specific response mediated by B and T lymphocytes that confers effective and long-lasting protection against infection. Adaptive immunity is based on the generation of a large repertoire of antigen-recognition receptors by somatic gene rearrangement. Diversity has been considered the hallmark of adaptive immunity. In the last few years, however, evidence has accumulated supporting the importance of diversity (probably as a response to selective pressures) even within innate immunity (Hoffmann et al. 1999). Moreover, differences in innate immune mechanisms have been shown to be critical in host susceptibility to infection (Cooke and Hill 2001), making this an area of intense research. The interferon-induced Mx1 protein is one of the best studied determinants of innate immunity to viral infection. In 1962, Lindenmann showed that the inbred mouse strain A2G is resistant to doses of mouseadapted influenza virus that are lethal to other inbred strains (Lindenmann 1962). This was a particularly interesting observation because innate resistance in A2G mice was dependent on a single dominant locus, namedMx1, that was expressed in a variety of cell types ranging from macrophages to hepatocytes and was exquisitely specific for orthomyxoviruses. Subsequent studies showed that the specific resistance of Mx1 murine cells to influenza viruses is attributable to the IFN-induced protein Mx1, and that after virus infection, the Mx1 protein is rapidly expressed in the nuclei of cells in the area where virus replication occurs, thus blocking viral spread (Arnheiter et al. 1996). The presence of a natural resistance gene to influenza was intriguing because mice are not natural hosts for orthomyxoviruses. Soon it became clear thatMx1was the first member of a small gene family present in all vertebrate species from fish to men, and that the spectrum of antiviral activity was much larger than initially appreciated. However, the exact mechanism of action of Mx proteins is still a matter of debate, and it is not clear whether the antiviral activity of Mx is a luxurious accident of some undefined cellular function or has evolved to inhibit in each species a set of species-specific pathogens. In this issue, Watanabe’s group describes a series of Mx alleles in chicken breeds some of which encode proteins with antiviral activity against influenza and vesicular stomatitis virus (VSV) (Ko et al. 2002). These findings are interesting from several perspectives. First, they close the chain of observations initiated with the discovery of the innate resistance in A2G mice to a mouse-adapted influenza virus by demonstrating, as has been speculated to exist, the presence of an active Mx protein in a species that functions as a reservoir for the virus. Second, they highlight the versatility of the antiviral function of the Mx protein that is associated with a single Ser631Asn substitution present in about 50% of the breeds studied. Third, these findings also trigger important questions: Is there a physiological function for Mx proteins? Is there a common theme in the mechanism of action of Mx against viruses? What are the mechanisms that fashioned the Mx antiviral activity? Most species have one to three Mx protein isoforms with different antiviral activities and intracellular localization. The prototype protein, mouse Mx1, as well as other rodent isoforms including rat Mx1, accumulate in the nucleus. In contrast, other isoforms as well as Mx proteins of humans and most other species are localized in the cytoplasm. Expression of Mx proteins is stimulated by IFN / or by viral infection irrespective of their activity. In fact, Mx proteins present a wide range of antiviral activity: nuclear Mx1 proteins inhibit principally orthomyxoviruses; in contrast, the cytoplasmic isoforms inhibit RNA viruses of the most diverse families, whereas some isoforms lack detectable antiviral function (Table 1). What is the structural basis of Mx function? Mx proteins belong to a superfamily of proteins, the large GTPases, which includes the dynamins, the products encoded by the Drosophila shibire, the yeast vacuolar sorting protein Vps1p, and the GTP-binding protein from Arabidopsis thaliana (van der Bliek 1999). Members of this family are present in a variety of cell locations where they perform a range of functions including endocytosis, intracellular vesicle transport, and mitochondria distribution. These proteins present modular organization characterized by at least three distinct functional domains with varied degrees of sequence conservation underlying their ability to self-assemble into higher order structures that resemble rings and helical stacks of rings, as in dynamin and Mx proteins (van der Bliek 1999). In particular, Mx proteins present a highly conserved N-terminal GTPase domain of ∼300 amino acids, a “middle” domain of ∼150 amino acids, and a GTPase effector domain (GED) of ∼100 amino acids including two leucine zippers that have the capacity to form amphipathic -helices (Fig. 1A). Studies with human MxA showed that the GED is able to specifically contact the middle domain, and that this interaction is critical to constitute a functional GTPase domain, as well as for oligomerization (Schumacher and Staeheli 1998; Di Paolo et al. 1999). Despite its functional and structural role that might be expected to restrict the amount of sequence variation among the different Mx proteins, the carboxy-terminal region shows 22% sequence identity as opposed to 38% identity in the remaining sequence (Fig. 1B). This would indicate that these sequence variations underlie functional differences between the different Mx isoforms as well as with other members of the dynamin family. Indeed, divergent sequences in the carboxy-terminus on other members of the dynamin family are thought to control specific localization and functional Corresponding author. E-MAIL svidal@uottawa.ca; FAX (613) 5625452. Article and publication are at http://www.genome. org/cgi/doi/10.1101/gr.20102. Insight/Outlook