Research on a Minor Organism can also be Benefit the World: The Fascinating Cellular Slime Mold Dictyostelium discoideum

Abstract

In 1985, when I entered the Graduate School of Science at Kyoto University, I began my research on cellular slime molds, a group of soil microorganisms. The cellular slime mold Dictyostelium discoideum is studied globally as a model organism for cell and developmental biology. I was conducting basic biological research into cell differentiation and migration using D. discoideum, and during this process, our research group made a discovery with potential implications for drug development. Specifically, we found that a chlorinated polyketide named differentiation-inducing factor 1 (DIF-1), derived from D. discoideum, exhibits antitumor activity. Based on this discovery, I began elucidating the mechanism of the antitumor action of DIF-1 and developing anticancer drugs using DIF-1 as a lead compound. During this period, in 1991, I obtained my Ph.D. in research related to D. discoideum cell differentiation, and subsequently served as a Japan Society for the Promotion of Science (JSPS) Special Research Fellow before joining the Institute for Molecular and Cellular Regulation (IMCR) at Gunma University in 1993. I then joined the Graduate School of Health and Sports Sciences at Juntendo University in 2015, where I have been until 2024. Throughout this period, I continued my research on DIF-1 and discovered that DIF-1 and its derivatives possess various biological activities ─ such as anti-diabetic, immunoregulatory, anti-bacterial, and anti-malarial activities ─ that could be applicable in drug development. In this review, I aim to present a segment of both our fundamental and applied research on D. discoideum and DIF-1.

Keywords: DIF-1; Dictyostelium discoideum; cellular slime mold; drug development.

Figure 1

(A) Fruiting bodies of the cellular slime mold Dictyostelium discoideum, each consisting of spores and a multicellular stalk. (B) Chemical structures of DIF-1 and differanisole A (DA). DIF-1, originally discovered in D. discoideum, acts as a stalk cell differentiation-inducing factor, while DA is an antitumor agent found in a fungal strain of the Chaetomium genus. (C) Schematic diagram illustrating how DIF-1 induces stalk cell differentiation (revised figure from ref. 12). DIF-1 may induce stalk cell differentiation by increasing [Ca²⁺]_i and [H⁺]_i possibly through a putative DIF-receptor in D. discoideum cells. Co-administration of Tg and DMO can replicate the effects of DIF-1 and induce stalk cell differentiation in the absence of DIF-1.

Figure 2

Proposed scheme for the antitumor effects of DIFs (revised figure from ref. 8). DIFs at concentrations ranging from several micromolars to several tens of micromolars trigger the indicated intracellular phenomena, leading to cell cycle arrest at G1/G0, induction/promotion of cell differentiation, and inhibition of cell migration^{8, 16-27)}. Notably, at appropriate concentrations, DIFs have inhibited cell growth in all tested tumor cell lines both in vitro and in vivo, and at higher concentrations, they have induced caspase-independent cell death in vitro^{18, 21, 25)}. DIFs also induce re-differentiation (erythroid differentiation) into hemoglobin-producing cells in murine and human leukemia (B8 and K562) cells^{16, 20)} and promote retinoic acid-induced granulocyte differentiation in human leukemia HL-60 cells in vitro¹⁹⁾. Additionally, DIFs suppress the migration and metastasis of certain cancer cells both in vitro and in vivo^{24, 26, 27)}. Chemical structure-activity relationship (SAR) analyses have revealed that several DIFs, such as DIF-3(+1) and Bu-DIF-3, show promise as lead compounds for the development of anticancer drugs^{8, 17)}. Abbreviations: ROS, reactive oxygen species; PAK1, p21-activated kinase 1; PDE1, calmodulin-dependent cAMP/cGMP phosphodiesterase; Erk, extracellular signal-regulated kinase; STAT3, signal transducer and activator of transcription 3; PI3K, phosphatidylinositol 3-kinase; GSK-3β, glycogen synthase kinase-3β; pRB, retinoblastoma protein.

Figure 3

Proposed scheme for the glucose uptake-promoting effect of DIF-1 (revised figure from ref. 33). Our findings have demonstrated that DIF-1 at concentrations of 10-20 μM promotes glucose uptake, at least in part, by uncoupling mitochondrial activity and inhibiting mitochondrial malate dehydrogenase (MDH2) activity, thereby reducing ATP production and activating AMP kinase (AMPK) in mammalian cells^{8, 28, 31, 33)}. It has also been suggested that DIF-1 promotes glucose uptake partly through an increase in cellular cAMP levels³²⁾. SAR analyses have revealed that both DIF-1 and DIF-1(3M) show promise as lead compounds for the development of anti-diabetes and anti-obesity drugs^{8, 28, 30)}.

Figure 4

Proposed scheme for the immune-regulatory effects of DIFs on ConA-induced IL-2 production in Jurkat T cells (revised figure from ref. 37). The mitogen ConA stimulates IL-2 production via TCR and subsequent activation (phosphorylation or dephosphorylation) of the MAPK/AP-1, IKK/NFκB, and CN/NFAT pathways in T-lymphocytes, whereas the immunosuppressive drugs cyclosporin A (CsA) and FK506 inhibit CN via the immunophilins^52-55). DIFs at several micromolar levels affect ConA-induced IL-2 production in vitro in Jurkat T cells. For example, Bu-DIF-3 and CP-DIF-3 suppress IL-2 expression by inhibiting NFAT and NFκB activities, whereas TH-DIF-1 and TM-DIF-1 suppress it, at least in part, by inhibiting NFAT and AP-1 activities³⁶⁾. In contrast, DIF-1(+1) and DIF-3(3M) promote ConA-induced IL-2 mRNA expression, at least in part, by enhancing AP-1 activity³⁷⁾. Abbreviations: ConA, concanavalin A; TCR, T-cell receptor; PLC, phospholipase C; IP3, inositol 1,4,5-trisphosphate; CN, calcineurin; DAG, diacylglycerol; PKC, protein kinase C; IKK, IκB kinase; MAPK, mitogen-activated protein kinase; NFAT, nuclear factor of activated T-cell; NFκB, nuclear factor kappa B; AP-1, activator protein-1, a transcription factor formed from a heterodimer of c-Fos and c-Jun. P- denotes a phosphate group.

Figure 5

(A) Chemical structures of several DIFs that have antimicrobial activities. DIF-3(+1) and Bu-DIF-3 suppress the infection and growth of T. cruzi in vitro with IC₅₀ values of 0.5-3 μM. Ph-DIF-3 and CP-DIF-3 suppress the growth of Gram-positive bacteria, including MRSA and VREs, in vitro with MIC (minimum inhibitory concentration) values of 1-3 μM. On the other hand, DIF-1(+2) and DIF-1(+3) suppress the growth of P. falciparum, including ART-resistant strains, in vitro with IC₅₀ values of 0.1-1.5 μM^, . (B) Proposed scheme for the anti-meiotic effect of DIF-1 in Xenopus oocytes. Progesterone induces germinal vesicle breakdown (GVBD), at least in part, by stimulating c-Mos (MAPKKK) production and subsequent activation of MAPKK and MAPK. DIF-1 at 20 μM inhibits progesterone-induced GVBD by targeting upstream of c-Mos production. Abbreviations: MAPK, mitogen-activated protein kinase; MAPKK, MAPK kinase; MAPKKK, MAPKK kinase. (C) Chemical structure of Br-DIF-1. Br-DIF-1 at 0.3-3 μM dose-dependently promotes DMSO-induced cardiomyocyte differentiation in mouse P19CL6 embryonic carcinoma cells.