No abstract

Polyamines are small, positively charged molecules derived from ornithine and synthesized through an intricately regulated enzymatic pathway. Within cells, they are abundant and play several roles in diverse processes. We find that polyamines are required for the life cycle of the RNA viruses chikungunya virus (CHIKV) and Zika virus (ZIKV). Depletion of spermidine and spermine via type I interferon signaling-mediated induction of spermidine/spermine N1-acetyltransferase (SAT1), a key catabolic enzyme in the polyamine pathway, restricts CHIKV and ZIKV replication. Polyamine depletion restricts these viruses in vitro and in vivo, due to impairment of viral translation and RNA replication. The restriction is released by exogenous replenishment of polyamines, further supporting a role for these molecules in virus replication. Thus, SAT1 and, more broadly, polyamine depletion restrict viral replication and suggest promising avenues for antiviral therapies.

Alphaviruses, such as chikungunya virus (CHIKV) and Sindbis virus (SINV), are vector-borne pathogens that cause acute illnesses in humans and are sometimes associated with neuropathies, especially in infants and elderly patients. Little is known about their mechanism of entry into the central nervous system (CNS), even for SINV, which has been used extensively as a model for viral encephalopathies. We previously established a CHIKV infection model in the optically transparent zebrafish larva; here we describe a new SINV infection model in this host. We imaged in vivo the onset and progression of the infection caused by intravenous SINV inoculation. Similar to that described for CHIKV, infection in the periphery was detected early and was transient, whereas CNS infection started at later time points and was persistent or progressive. We then tested the possible mechanisms of neuroinvasion by CHIKV and SINV. Neither virus relied on macrophage-mediated transport to access the CNS. CHIKV, but not SINV, always infects endothelial cells of the brain vasculature. By contrast, axonal transport was much more efficient with SINV than CHIKV, both from the periphery to the CNS and between neural tissues. Thus, the preferred mechanisms of neuroinvasion by these two related viruses are distinct, providing a powerful imaging-friendly system to compare mechanisms and prevention methods of encephalopathies.

Polyamines, which are small positively charge molecules present in all cells, play important roles in the replication of DNA and RNA viruses. Chikungunya virus (CHIKV) relies on polyamines for translation of the viral genome upon viral entry, and pharmacological depletion of polyamines limits viral replication. However, the potential development of antiviral resistance necessitates a better understanding of how polyamines function and can be targeted via compounds that alter polyamine levels. We have isolated CHIKV that is resistant to polyamine depletion and contains two mutations in the nonstructural protein 1 (nsP1)-coding region in combination with a mutation to the opal stop codon preceding nsP4. These mutations, in addition to promoting viral replication in polyamine-depleted cells, confer enhanced viral replication in vitro and in vivo. The nsP1 mutations enhance membrane binding and methyltransferase activities, while the stop codon mutation allows increased downstream translation. These mutations, when combined, enhance viral fitness, but individual mutants are attenuated in mosquitoes. Together, our results suggest that CHIKV can evolve resistance to polyamine depletion and that pharmaceuticals targeting the polyamine biosynthetic pathway may be best used in combination with other established antivirals to mitigate the development of resistance.IMPORTANCE Chikungunya virus is a mosquito-borne virus that has infected millions worldwide. Its expansion into the Americas and rapid adaptation to new mosquito hosts present a serious threat to human health, which we can combat with the development of antiviral therapies as well as understanding how these viruses will mutate when exposed to antiviral therapies. Targeting polyamines, small positively charged molecules in the cell, may be a potential strategy against RNA viruses, including chikungunya virus. Here, we have described a virus that is resistant to polyamine depletion and has increased fitness in cells and in full organisms. Mutations in viral genome capping machinery, membrane binding activity, and a stop codon arise, and their altered activities enhance replication in the absence of polyamines. These results highlight strategies by which chikungunya virus can overcome polyamine depletion and emphasize continued research on developing improved antiviral therapies.KEYWORDS antiviral, antiviral resistance, chikungunya virus, polyamines, viral replication C hikungunya virus (CHIKV) is an arthropod-borne virus (arbovirus) from the Alphavirus family that has caused several severe outbreaks worldwide. Clinical manifestations can be severe and long lasting, including fever and chronic joint pain, and rapid