## 4 Materials and Methods
### Statistics and Data Visualisation  
Graphs were generated and tests of statistical significance were performed using Prism (version 6.01; GraphPad Software, La Jolla, CA, USA). Statistical significance was calculated by multiple unpaired \(t\)-tests of at least 3 biological replicates. \(p\)-values \(\leq\) 0.01 were considered as being statistically significant.  
Extracellular data are presented as relative concentration difference \(\Delta[\mathrm{c}]\) between the indicated time point \(\mathrm{t_{x}}\) and \(\mathrm{t_{0}}\), normalized to changes in cell number (cell number \(\mathrm{t_{x}}\)/cell number \(\mathrm{t_{0}}\)). Values are presented as bar charts or colour coded charts, which were created using MeV v4.8.1 [64] with the following settings for hierarchical clustering: optimized gene leaf order, euclidean distance metric and average linkage method.  
Intracellular data are presented as volcano plots and word clouds [65], colour coded charts or bar charts. For relative concentrations, computed values of the signal area were normalized on the corresponding internal standard and the cell number. For calculation of fold changes, missing values were replaced with half the value of the lowest abundant metabolite that was analysed (Supplementary Material) and assumed to be the detection limit.  
**Supplementary Materials:** The following are available online at [[http://www.mdpi.com/2218-1989/6/4/41/s1](http://www.mdpi.com/2218-1989/6/4/41/s1)]([http://www.mdpi.com/2218-1989/6/4/41/s1](http://www.mdpi.com/2218-1989/6/4/41/s1)), Figure S1. (A) Uptake and secretion of amino acids by A549 cells after 12 h of cytochalasin D treatment. Extracellular concentrations changes of amino acids were normalized on the proliferation rate (\([\mathrm{t_{cn}}\)-\(\mathrm{c_{0}}]\))/(cell number\(\mathrm{t_{0}}\)). Data are presented as mean (\(n=5\)) in a colour coded chart. Red fields indicate an increase in the extracellular concentration and blue fields indicate a decrease of the metabolite concentration compared to the initial concentration. (B) Mean percentages of incorporation levels of D-glucose-\({}^{13}\)C\({}_{6}\) of A549 cells without and with 12 h treatment with cytochalasin D are presented (\(n=4\)). Proportions of complete labelled nucleotides and nucleotide-sugars are black, labelled sugar moieties are presented as grey pattern and unlabelled metabolite amounts are displayed as light grey; Table S1: extracellular data, Table S2: intracellular data (infection); Table S3: intracellular data (metabolic inhibitor assay); Table S4: intracellular data (metabolic labelling assay); Table S5: molecular masses of identified metabolites.  
**Acknowledgments:** This study was supported by funds from the Bundesministerium fur Bildung und Forschung (BMBF) ([[http://www.unternehmen-region.de/](http://www.unternehmen-region.de/)]([http://www.unternehmen-region.de/](http://www.unternehmen-region.de/))) to [PERSON] and [PERSON] within the framework of ZIKV-FunGene (03Z1 CN21) and to [PERSON], [PERSON] and [PERSON] within the framework Infect-ERA \"EUGENPATH\" (031A410B). It was furthermore supported by funds from the Deutsche Forschungsgemeinschaft ([[http://www.dfg.de/](http://www.dfg.de/)]([http://www.dfg.de/](http://www.dfg.de/))) to [PERSON], [PERSON] and [PERSON] within the framework SFB/TRR34 (Z4) and RTG1870. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Moreover, we are thankful to [PERSON] and [PERSON] for helpful discussions and critical reading of the manuscript and to Baltic Analytics GmbH for sharing laboratory facilities.  
**Author Contributions:** [PERSON] and [PERSON] conceived and designed the experiments; [PERSON] and [PERSON] performed the experiments; [PERSON] and [PERSON] analysed the data; [PERSON] and [PERSON] contributed reagents/materials/analysis tools; and [PERSON] and [PERSON] wrote the paper.  
**Conflicts of Interest:** The authors declare no conflict of interest.