Abstract
Many viruses depend on host microtubule motors to reach their destined intracellular location. Viral particles of neurotropic alphaherpesviruses such as herpes simplex virus 1 (HSV1) show bidirectional transport towards the cell center as well as the periphery, indicating that they utilize microtubule motors of opposing directionality. To understand the mechanisms of specific motor recruitment, it is necessary to characterize the molecular composition of such motile viral structures. We have generated HSV1 capsids with different surface features without impairing their overall architecture, and show that in a mammalian cell-free system the microtubule motors dynein and kinesin-1 and the dynein cofactor dynactin could interact directly with capsids independent of other host factors. The capsid composition and surface was analyzed with respect to 23 structural proteins that are potentially exposed to the cytosol during virus assembly or cell entry. Many of these proteins belong to the tegument, the hallmark of all herpesviruses located between the capsid and the viral envelope. Using immunoblots, quantitative mass spectrometry and quantitative immunoelectron microscopy, we show that capsids exposing inner tegument proteins such as pUS3, pUL36, pUL37, ICP0, pUL14, pUL16, and pUL21 recruited dynein, dynactin, kinesin-1 and kinesin-2. In contrast, neither untegumented capsids exposing VP5, VP26, pUL17 and pUL25 nor capsids covered by outer tegument proteins such as vhs, pUL11, ICP4, ICP34.5, VP11/12, VP13/14, VP16, VP22 or pUS11 bound microtubule motors. Our data suggest that HSV1 uses different structural features of the inner tegument to recruit dynein or kinesin-1. Individual capsids simultaneously accommodated motors of opposing directionality as well as several copies of the same motor. Thus, these associated motors either engage in a tug-of-war or their activities are coordinately regulated to achieve net transport either to the nucleus during cell entry or to cytoplasmic membranes for envelopment during assembly.
Highlights
To reach their destined subcellular location, viruses utilize motor proteins that move unidirectional along microtubules (MT) or actin filaments [1,2,3,4,5]
We established a cell-free system that reconstitutes the binding of native, mammalian microtubule motors to intact tegumented herpes simplex virus (HSV) capsids
Our data suggest that the inbound motor dynein and the outbound motor kinesin-1 bind directly and independently of other host factors to the inner tegument that coats the capsids during cytosolic transport
Summary
To reach their destined subcellular location, viruses utilize motor proteins that move unidirectional along microtubules (MT) or actin filaments [1,2,3,4,5]. Digital time-lapse microscopy experiments in living cells have provided fundamental insights into the dynamics of intracellular transport, and demonstrated that host and viral cargos show rapid reversals in transport direction. This raises the question how net transport towards a particular destination is achieved and regulated. The cargo surface could either engage exclusively inbound or outbound motors according to the hypothesis of ‘‘exclusive presence’’, or the capsids could recruit motors of opposing directionality simultaneously [6,7,8,9]. The hypothesis of ‘‘coordinated regulation’’ proposes that the activity of the bound motors is coordinately regulated so that they do not interfere with each other [13,14,15]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
