Fig. 5: TFEB promotes non-lytic release of CVB3 in early stages of infection.

a Accumulation of extracellular virus is delayed in cells lacking TFEB. Wild-type (HAP1^WT) and TFEB knockout (TFEB^KO) HAP1 cells were infected with CVB3 at MOI 1 as in Fig. 4a. Intracellular and extracellular virus titers at the indicated time points were determined by end-point titration. Shown are means, error bars represent ± SEM (n = 3 independent experiment, each with biological triplicates; results of all replicates were averaged). Results were statistically evaluated by two-way ANOVA with Bonferroni’s post-hoc test. ***, p < 0.001 (p = 0.0005). Plots showing the individual data points are available in the Source data file. b Cell viability at 8 hpi is not affected by TFEB knockout. HAP1^WT or TFEB^KO HAP1 cells were infected with CVB3 at MOI 10. Cell viability at 8 hpi was determined using a FACS-based viability staining. Shown are means, error bars represent ± SD. Different symbols indicate data of three independent experiments. c TFEB knockout impairs extracellular vesicle-enclosed virus release. Virus released in the supernatant at 8 hpi was isolated by pelleting at 100,000 × g and purified using density gradient centrifugation. Infectivity retrieved within naked virus (1.35–1.15 g ml⁻¹) and EV-enclosed virus containing fractions (1.10–1.04 g ml⁻¹) was determined using end-point titration. The ratio between virus recovery from samples from HAP1^WT and TFEB^KO HAP1 cells is depicted after correction for any difference in intracellular titers. Shown are means, error bars represent ± SD (n = 3 independent experiments). Results were statistically evaluated using a one-sample t-test. Details on the statistics are included in the Source data file. **, p < 0.005 (p = 0.0017). d, e TFEB knockout affects the induction of secretory autophagy. Western blot analysis of the autophagy-related protein LC3 in whole cell lysates (d) and 100,000 × g pelleted material from cell culture supernatants (e) from CVB3-infected HAP1^WT or TFEB^KO HAP1 cells at 8 hpi. Representative image of three independent experiments are shown. Source data are provided as a Source Data file for all panels.