Fig 2: PTS was highly expressed in LUAD. (A) PTS expression level in LUAD in TCGA data. (B) PTS expression level in LUAD based on individual cancer stages. (C) Correlation analysis of PTS level and prognosis of LUAD patients. Expression of PTS level in LUAD tissues was measured by Western blot (D) and immunohistochemistry (stained with PTS antibody; magnification 40×) (E). PTS mRNA and protein level in LUAD cells was tested by qRT-PCR (F) and Western blot (G). **, P<0.01, compared with the indicated group. TCGA, The Cancer Genome Atlas; LUAD, lung adenocarcinoma; mRNA, messenger RNA; PTS, 6-pyruvoyl-tetrahydropterin synthase; HR, hazard ratio; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; qRT-PCR, quantitative real-time polymerase chain reaction.

Fig 3: Silencing of PTS inhibited LUAD cell migration and invasion. (A) Cell migration was tested by wound healing assay (magnification 4×). Transwell assay was used to test cell migration (B) (stained with crystal violet; magnification 10×) and invasion (C) (stained with crystal violet; magnification 10×). (D) Levels of E-cadherin, vimentin and N-cadherin were tested by Western blot. **, P<0.01, compared with the indicated group. si-NC, siRNA-negative control; siRNA, small interfering RNA; si-PTS, siRNA for PTS; PTS, 6-pyruvoyl-tetrahydropterin synthase; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; LUAD, lung adenocarcinoma.

Fig 4: ATF4 is an upstream transcription factor of PTS. (A) PTS upstream transcription factors were predicted by HumanTFDB. (B) Linear analysis of PTS and ATF4. (C) Interaction between ATF4 and PTS was verified by dual luciferase assay. ATF4 mRNA (D) and PTS mRNA (E) were tested by qRT-PCR. Cell proliferation, apoptosis, and invasion were tested by colony formation assay (stained with crystal violet; magnification 100×) (F), flow cytometry (G), and transwell assay (stained with crystal violet; magnification 200×) (H). #, P<0.05, ##, P<0.05, compared with the si-PTS-1 group. *, P<0.05; **, P<0.01, compared with the si-NC group. PTS, 6-pyruvoyl-tetrahydropterin synthase; ATF4, activating transcription factor 4; CBX3, chromobox 3; HDAC2, histone deacetylase 2; NFYB, nuclear transcription factor Y subunit beta; EZH2, enhancer of zeste 2 polycomb repressive complex 2 subunit; RNF2, ring finger protein 2; ZNF143, zinc finger protein 143; NC, negative control; WT, wild type; Mut, mutant; mRNA, messenger RNA; si-NC, siRNA-negative control; siRNA, small interfering RNA; si-PTS, siRNA for PTS; qRT-PCR, quantitative real-time polymerase chain reaction.

Fig 5: Silencing of PTS inhibited LUAD cell viability and proliferation and accelerated apoptosis. (A) PTS protein level was tested by Western blot. Cell viability and apoptosis were determined by CCK-8 (B), colony formation assay (stained with crystal violet; magnification 100×) (C), and flow cytometry (D), respectively. (E) Levels of Bcl-2, PARP1, and Bax were tested by Western blot. *, P<0.05; **, P<0.01, compared with the si-NC group or vector group. PTS, 6-pyruvoyl-tetrahydropterin synthase; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; si-NC, siRNA-negative control; siRNA, small interfering RNA; si-PTS, siRNA for PTS; OD, optical density; PARP1, poly-ADP-ribose polymerase; Bax, Bcl-2-associated X protein; Bcl-2, B-cell lymphoma 2; LUAD, lung adenocarcinoma; CCK-8, Cell Counting Kit-8.