The research, published in the open-access journal MicrobiologyOpen, is notable for being the first comprehensive characterization of the pigment chemistry of M. terreus, a species only described in 2010 and rarely studied since. While its better-known relative M. luteus has been investigated extensively for biotechnologically useful pigments such as beta-carotene and zeaxanthin, the chemical composition and biological activities of M. terreus pigments had remained essentially unexplored. The new study set out to close three gaps at once: establishing what pigments the bacterium actually makes, testing those pigments against cancer types not previously examined, and probing the molecular mechanisms behind any anticancer effects using computational tools.

The identification of the isolate began conventionally. Soil samples from poplar forests, pine forests, potting soil, and agricultural fields were diluted, plated on tryptic soy agar, and incubated at several temperatures. Colonies showing distinct yellow, red, or pink pigmentation were purified, and fifteen distinct isolates were screened quantitatively for pigment production in liquid culture. SK34, recovered from pine forest soil, produced the highest total pigment content. Sequencing of its 16S ribosomal RNA gene and comparison with GenBank records placed it firmly within the Micrococcus group, with its closest relative being M. terreus DSM 28110. The sequence has been deposited in GenBank under accession number PX765938.

The chemical heart of the study came from liquid chromatography coupled to quadrupole time-of-flight mass spectrometry, a technique that separates compounds and measures their accurate masses to within five parts per million. Because carotenoids are easily degraded by light and heat, all analyses were performed under low-light conditions at controlled temperatures. The analysis revealed eight distinct carotenoids in the extract. Phytoene dominated at nearly 31 percent of the total, followed by zeaxanthin at 17.1 percent, beta-cryptoxanthin at 13.7 percent, and beta-carotene at 10.4 percent, with smaller amounts of astaxanthin, alpha-carotene, lycopene, and sarcinaxanthin. Since phytoene is colorless, the researchers concluded that the abundant yellow zeaxanthin is the main contributor to the bacterium’s vivid pigmentation.

Biological testing showed the extract was active against all five cancer lines examined: liver (HepG2), lung (A549), neuroblastoma (SH-SY5Y), breast (MCF-7), and cervical (HeLa) cells, with effects that increased in a dose-dependent manner. The concentration needed to halve cell viability after 24 hours was lowest for SH-SY5Y cells at 61.91 micrograms per milliliter, followed by MCF-7 at 82.08, A549 at 97.60, HeLa at 153.74, and HepG2 at 243.76 micrograms per milliliter. Crucially, healthy human dermal fibroblasts retained more than 90 percent viability even at the highest concentration tested, 1000 micrograms per milliliter. Because no inhibitory concentration could be established for the healthy cells within that range, the researchers reported minimum selectivity indices, which exceeded 16 for SH-SY5Y, 12 for MCF-7, and 10 for A549. Values above 2 are generally considered indicative of selective cytotoxicity, and values above 10 reflect high selectivity, placing this extract in promising territory.

To understand how the extract kills cancer cells, the team turned to the two cellular self-destruction programs most relevant to cancer biology: apoptosis, or programmed cell death, and autophagy, the cellular recycling process that can tip into lethal overdrive under stress. Apoptosis is governed by a delicate balance between pro-death signals such as p53, Bax, and the caspase enzymes, and anti-death proteins such as Bcl-2 and Akt1. Cancer cells frequently disable these death circuits, so therapies that restore them are a major research focus. The researchers treated SH-SY5Y cells at the extract’s inhibitory concentration and measured gene expression by quantitative real-time PCR.

The gene expression results were unambiguous. Expression of p53, Bax, caspase-3, caspase-8, and caspase-9 all rose significantly, while the antiapoptotic genes Bcl-2 and Akt1 were suppressed. The ratio of Bax to Bcl-2, a widely used molecular indicator of apoptotic commitment, increased markedly in treated cells. Notably, caspase-9, the initiator caspase of the intrinsic or mitochondrial pathway, showed the largest change, suggesting the extract primarily activates the internal suicide program rather than the external death-receptor route, although the concurrent rise in caspase-8 hints that both pathways may be engaged. Autophagy genes Beclin-1 and mTOR shifted only slightly and not significantly, indicating that apoptosis, not autophagy, is the dominant mechanism.

Protein-level measurements corroborated the transcript data. Enzyme-linked immunosorbent assays showed significantly elevated Bax protein, reduced Bcl-2, a sharply increased Bax/Bcl-2 ratio, and higher levels of cleaved caspase-3 and cleaved caspase-9, with a smaller but significant rise in cleaved caspase-8. Flow cytometry using Annexin V and propidium iodide staining provided the final piece of evidence: in untreated cultures, 92.32 percent of SH-SY5Y cells were viable, but after extract treatment the viable fraction fell to 71.45 percent, while early apoptotic cells rose to 14.13 percent and late apoptotic cells to 10.99 percent, for a total apoptosis rate of 25.12 percent. This orderly progression from early to late apoptosis is exactly the pattern desired of a candidate anticancer agent.

Computational docking added a mechanistic hypothesis about which individual carotenoids matter most. The five major compounds were docked against three apoptosis-related proteins: Akt1, p53, and Bcl-2. Against Akt1, beta-carotene bound most strongly at minus 9.0 kilocalories per mole, outperforming even the reference drug capivasertib. Against Bcl-2, zeaxanthin led at minus 9.1, with astaxanthin close behind. Against p53, astaxanthin achieved minus 7.8 kilocalories per mole. As expected for long, lipophilic polyene chains, the carotenoids relied overwhelmingly on hydrophobic and van der Waals contacts within the binding pockets, engaging conserved residues such as phenylalanine, tyrosine, and valine. The authors caution that docking predicts thermodynamic feasibility of binding rather than functional activation, and that molecular dynamics simulations would be needed to confirm stability. Still, the combined evidence suggests the extract’s components may simultaneously inhibit cell-survival proteins such as Bcl-2 and Akt1 while supporting pro-apoptotic signaling through p53 and the caspase cascade.

Perhaps the most intriguing implication is that no single carotenoid can claim credit. The researchers argue that the coexistence of eight carotenoids may generate additive or synergistic effects, since mixtures can modulate multiple molecular targets at once, an advantage in cancer therapy where coordinated pathway modulation generally outperforms single-target approaches. The team also acknowledges limitations: the mechanistic analyses used the inhibitory concentration, so some observed changes may be downstream consequences of apoptosis rather than its earliest triggers, and time-course studies at lower doses will help disentangle cause from effect. In vivo validation remains the essential next step before any clinical relevance can be claimed. Nevertheless, the work transforms an obscure soil bacterium into a credible source of bioactive pigments and illustrates how integrated metabolomics, cell biology, and computational chemistry can accelerate the search for natural anticancer compounds hiding in ordinary dirt.

Subject of Research: Anticancer activity of a carotenoid-rich pigment extract from Micrococcus terreus SK34 against human cancer cell lines

Article Title: Carotenoid‐Rich Pigment Extract From Micrococcus terreus SK34 Induces Apoptosis in SH‐SY5Y Neuroblastoma Cells: Integrated Metabolomic, Molecular Docking, and Experimental Validation

Article References: Karaagac, S. I., Albayrak, S., ShadiDizaji, A., Kiziler, M. E., Eroglu, E., Arslan, N. P., & Taskin, M. (2026). Carotenoid‐Rich Pigment Extract From Micrococcus terreus SK34 Induces Apoptosis in SH‐SY5Y Neuroblastoma Cells: Integrated Metabolomic, Molecular Docking, and Experimental Validation. MicrobiologyOpen, 15(5), Article e70382. https://doi.org/10.1002/mbo3.70382

Image Credits: AI Generated

DOI: 10.1002/mbo3.70382

Keywords: carotenoids, Micrococcus terreus, neuroblastoma, apoptosis, SH-SY5Y, molecular docking, LC-Q-TOF-MS, natural pigments, anticancer, zeaxanthin, Bcl-2, soil bacteria