The methionine and arginine biosynthetic genes are expressed in response to neutrophils and require fresh human serum. Columns 1–7 are the time course of Saccharomyces and neutrophils cultured in RPMI medium 1640 containing 10% fresh human serum for 10 (column 1), 20 (column 2), 40 (columns 3 and 4), 60 (columns 5 and 6), or 80 (column 7) minutes. Columns 8 and 9 are the results of the heat-inactivated serum experiment, in which heat-inactivated human serum (column 9) or fresh human serum (column 8) was included in the medium. Columns 10–12 are the results of a filter experiment, in which a filter was used to separate yeast from neutrophils. In column 10 yeast and neutrophils were on the same side of the filter. In column 11 yeast was on one side of the filter and neutrophils were on the other. In column 12 neutrophils and yeast were on one side and yeast was on the other (the yeast alone was assayed). All 83 genes that met the criteria for induction were selected for illustration purposes. Genes were divided into four groups: methionine (M), arginine (R), other amino acid genes (aa), and other genes (o). ADE and URA genes served as control (*) genes. Results are visualized by using cluster and treeview.

In Saccharomyces the response to ingestion by neutrophils is similar to that engendered by amino acid deprivation. (A) Saccharomyces was cultured with neutrophils in RPMI medium 1640 containing 10% fresh autologous human serum (columns 4–8) or in minimal medium without neutrophils (columns 1–3) for 40 min (columns 1–5 and 8) or 60 min (columns 6 and 7). Yeast strains were wild type (columns 2–8) or gcn4Δ (column 1). Strain backgrounds were EM93 (columns 3–7) or S288C (columns 1, 2, and 8). For purposes of illustration, amino acid and nucleic acid biosynthetic genes were selected for cluster analysis. ADE and URA genes served as control genes (*). Experiments were clustered by using cluster and treeview. (B) The transcriptional profile in response to neutrophils (N) was compared with amino acid (aa) deprivation conditions.

The expression profile of yeast in response to neutrophils is not dominated by a response to oxidative stress. Yeast cells were cultured in the presence of 0.4 mM H₂O₂ (column 1) or with neutrophils (columns 2 and 3) for 40 min. All 83 genes meeting criteria for induction were selected for illustration purposes. Genes were divided into four groups: methionine (M), arginine (R), other amino acid genes (aa), and other genes (o). ADE and URA genes served as control (*) genes. Experiments were clustered by using cluster and treeview.

Response of Candida to ingestion by neutrophils. In Candida the response to ingestion by neutrophils is similar to that engendered by amino acid deprivation. (A) Candida was cultured with neutrophils in RPMI medium 1640 containing 10% fresh human serum for 60 (column 5) or 80 (column 4) min or in minimal medium for 40 (column 3), 60 (column 1), or 80 (column 2) min. Amino acid and nucleic acid biosynthetic genes were selected for cluster analysis for illustration purposes. ADE and URA genes served as controls (*). Experiments were clustered by using cluster and treeview. (B) Candida was cultured in minimal medium with (+) or without (–) amino acids (Upper) or in RPMI medium 1640 containing 10% fresh human serum with (+) or without (–) neutrophils (Lower) for 60 min. Candida mRNA was subjected to Northern analysis, using probes for the genes indicated. Ethidium bromide staining of ribosomal RNA to control for loading is shown in the lower gels. (C) Candida does not form filaments inside neutrophils. Yeast-form Candida cells were cultured with monocytes (Upper) or neutrophils (Lower) in RPMI medium 1640 containing 10% fresh human serum for 1 h. (Bar is 20 μm.) See also Fig. 2, supplementary data at http://jura.wi.mit.edu/fink_public/neutrophils/.

Yeast methionine and arginine genes are induced in neutrophils, but not in monocytes. Yeast cells were cultured with neutrophils (columns 3 and 4) or monocytes (columns 1 and 2) in RPMI medium 1640 containing 10% fresh human serum for 40 min. All 83 genes meeting criteria for induction in neutrophils were selected for illustration purposes. Genes were divided into four groups: methionine (M), arginine (R), other amino acid genes (aa), and other genes (o). ADE and URA genes served as control (*) genes. Experiments were clustered by using cluster and treeview.