Fig 2: The expression of S1P in hnRNP H1- or SGPL1-downregulated HCT116 cells as assessed by ELISA. ELISA (n = 3) showed that the intracellular S1P expression was not changed in hnRNP H1- or SGPL1-downregulated HCT116 cells. The error bars show the SD.

Fig 3: hnRNP H1 upregulated sphingosine-1-phosphate lyase 1 (SGPL1) mRNA and promoted colorectal cancer progression. (A) The SRB assay (n = 5) showed that SGPL1 downregulation resulted in the strongest inhibition of cell growth in HCT116 cells. (B) RNA immunoprecipitation combined with RT-PCR (n = 3) confirmed the direct binding of hnRNP H1 and SGPL1 mRNA. (C) RT-PCR (n = 3) and western blotting (n = 3) showed the decrease in SGPL1 mRNA and protein in hnRNP H1-downregulated HCT116 cells. (D) RT-PCR (n = 3) and western blotting (n = 3) showed that the downregulation of SGPL1 mRNA and protein was not observed in hnRNP H1-downregulated HECE-1CT cells. (E) The SRB assay (n = 5) showed that an additional tumor-suppressive effect was not observed by SGPL1 downregulation. (F) The SRB assay (n = 5) showed that cell growth was significantly increased in the hnRNP H1-downregulated HCT116 cells with S1P in a concentration-dependent manner compared to those without sphingosine-1-phosphate (S1P). The error bars show the SD. * p < 0.05 by Student’s t-test.

Fig 4: Treatment of newborn Sgpl1-KO mice with AAV-SPL prolongs survival.(A) Treatment schedule, with upper images showing injection site and lower image showing green color of properly injected KO pup (AAV) next to untreated WT pup (Cntrl). (B) Images showing size discrepancy between WT and untreated Sgpl1-KO littermates at 17 DOL versus similarity in size of WT and AAV-SPL–treated Sgpl1-KO (AAV) littermates at 22 DOL. (C) Weight gain of female WT (blue), untreated Sgpl1-KO (red), and AAV-SPL–treated KO (green) mice. (D) Kaplan-Meier survival curve for WT (blue), untreated Sgpl1-KO (red), AAV-SPL–treated KO (green), and AAV-SPLK353L–treated KO (black) mice. Log-rank test with Bonferroni’s correction: for WT vs. all other groups, P < 0.0003; AAV vs. KO, P < 0.0003; KO vs. K353L, no significant difference (NSD).

Fig 5: Fold expression of mitochondrial fusion (MFN1, MFN2) and fission (DNM1L) genes at mRNA and protein level in control and patient fibroblasts and HeLa cell lines. A. MFN1/2 levels are reduced in FIB_65fs whereas MFN2 appears upregulated in FIB_545del while DNM1L was downregulated in both patient fibroblasts. B. MFN1 levels are increased in SGPL1-KO cells whereas DNM1L (DRP1) appears significantly downregulated.C. Western blot showing levels of DRP1 protein expression with similar reduction seen in FIB_65fs but not FIB_545del. D. DRP1 protein expression with reduction consistent with mRNA levels seen in SGPL1-KO HeLa cell lysates. (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001) Data are presented as the mean of three experiments.