From ecology to oncology: To understand cancer stem cell dormancy, ask a Brine shrimp (Artemia).

The brine shrimp (Artemia), releases embryos that can remain dormant for up to a decade. Molecular and cellular level controlling factors of dormancy in Artemia are now being recognized or applied as active controllers of dormancy (quiescence) in cancers. Most notably, the epigenetic regulation by SET domain-containing protein 4 (SETD4), is revealed as highly conserved and the primary control factor governing the maintenance of cellular dormancy from Artemia embryonic cells to cancer stem cells (CSCs). Conversely, DEK, has recently emerged as the primary factor in the control of dormancy exit/reactivation, in both cases. The latter has been now successfully applied to the reactivation of quiescent CSCs, negating their resistance to therapy and leading to their subsequent destruction in mouse models of breast cancer, without recurrence or metastasis potential. In this review, we introduce the many mechanisms of dormancy from Artemia ecology that have been translated into cancer biology, and herald Artemia's arrival on the model organism stage. We show how Artemia studies have unlocked the mechanisms of the maintenance and termination of cellular dormancy. We then discuss how the antagonistic balance of SETD4 and DEK fundamentally controls chromatin structure and consequently governs CSCs function, chemo/radiotherapy resistance, and dormancy in cancers. Many key stages from transcription factors to small RNAs, tRNA trafficking, molecular chaperones, ion channels, and links with various pathways and aspects of signaling are also noted, all of which link studies in Artemia to those of cancer on a molecular and/or cellular level. We particularly emphasize that the application of such emerging factors as SETD4 and DEK may open new and clear avenues for the treatment for various human cancers.

Ciliary ectosomes: transmissions from the cell's antenna.

The cilium is the site of function for a variety of membrane receptors, enzymes and signal transduction modules crucial for a spectrum of cellular processes. Through targeted transport and selective gating mechanisms, the cell localizes specific proteins to the cilium that equip it for the role of sensory antenna. This capacity of the cilium to serve as a specialized compartment where specific proteins can be readily concentrated for sensory reception also makes it an ideal organelle to employ for the regulated emission of specific biological material and information. In this review we present and discuss an emerging body of evidence centered on ciliary ectosomes - bioactive vesicles released from the surface of the cilium.

Proteins of the ciliary axoneme are found on cytoplasmic membrane vesicles during growth of cilia.

The cilium is a specialized extension of the cell in which many specific proteins are admitted and retained, while many others are excluded or expelled. In order to maintain the organelle, the cell must possess mechanisms for the selective gating of protein entry, as well as for the targeted transport of proteins to the cilium from their sites of synthesis within the cell [1-4]. We hypothesized that the cell employs cytoplasmic vesicles as vehicles not only for the transport of proteins destined for the ciliary membrane but also for the transport of axonemal proteins to the cilium by means of peripheral association with vesicles. To test this hypothesis, we employed two different experimental strategies: (1) isolation and biochemical characterization of cytoplasmic vesicles that carry ciliary proteins, and (2) in situ localization of ciliary proteins on cytoplasmic vesicle surfaces using gold labeling and electron microscopy. Our findings indicate that structural proteins destined for the ciliary axoneme are attached to the outer surfaces of cytoplasmic vesicles that carry integral ciliary membrane proteins during the process of ciliary growth.

The cilium secretes bioactive ectosomes.

The release of membrane vesicles from the surface of cells into their surrounding environment is now recognized as an important pathway for the delivery of proteins to extracellular sites of biological function. Membrane vesicles of this kind, termed exosomes and ectosomes, are the result of active processes and have been shown to carry a wide array of biological effector molecules that can play roles in cell-to-cell communication and remodeling of the extracellular space. Degradation of the extracellular matrix (ECM) through the regulated release of proteolytic enzymes is a key process for development, morphogenesis, and cell migration in animal and plant cells. Here we show that the unicellular alga Chlamydomonas achieves the timely degradation of its mother cell wall, a type of ECM, through the budding of ectosomes directly from the membranes of its flagella. Using a combination of immunoelectron microscopy, immunofluorescence microscopy, and functional analysis, we demonstrate that these vesicles, which we term ciliary ectosomes, act as carriers of the proteolytic enzyme necessary for the liberation of daughter cells following mitosis. Chlamydomonas has proven to be the key unicellular model for the highly conserved mechanisms of mammalian cilia, and our results suggest that cilia may be an underappreciated source of bioactive, extracellular membrane vesicles.

IFT proteins accumulate during cell division and localize to the cleavage furrow in Chlamydomonas.

Intraflagellar transport (IFT) proteins are well established as conserved mediators of flagellum/cilium assembly and disassembly. However, data has begun to accumulate in support of IFT protein involvement in other processes elsewhere in the cell. Here, we used synchronous cultures of Chlamydomonas to investigate the temporal patterns of accumulation and localization of IFT proteins during the cell cycle. Their mRNAs showed periodic expression that peaked during S and M phase (S/M). Unlike most proteins that are synthesized continuously during G1 phase, IFT27 and IFT46 levels were found to increase only during S/M phase. During cell division, IFT27, IFT46, IFT72, and IFT139 re-localized from the flagella and basal bodies to the cleavage furrow. IFT27 was further shown to be associated with membrane vesicles in this region. This localization pattern suggests a role for IFT in cell division.

Targeted gene disruption of dynein heavy chain 7 of Tetrahymena thermophila results in altered ciliary waveform and reduced swim speed.

Tetrahymena thermophila swims by the coordinated beating of hundreds of cilia that cover its body. It has been proposed that the outer arm dyneins of the ciliary axoneme control beat frequency, whereas the inner arm dyneins control waveform. To test the role of one of these inner arms, dynein heavy chain 7 protein (Dyh7p), a knockout mutant was generated by targeted biolistic transformation of the vegetative macronucleus. Disruption of DYH7, the gene which encodes Dyh7p, was confirmed by PCR examination of both genomic and cDNA templates. Both intact and detergent extracted, reactivated cell model preparations of these mutants, which we call DYH7neo3, displayed swim speeds that were almost half that of wild-type cells. Although the DYH7neo3 mutants were slower than wild type, they were able to modulate their swim speed and show ciliary reversal in response to depolarizing stimuli. High-speed video microscopy of intact, free-swimming DYH7neo3 mutants revealed an irregular pattern of ciliary beat and waveform. The mutant cilia appeared to be engaging in less coordinated, swiveling movements in which the typical shape, periodicity and coordination seen in wild-type cilia were absent or disturbed. We propose that the axonemal inner arm dynein heavy chain 7 proteins contribute to the formation of normal ciliary waveform, which in turn governs the forward swimming velocity of these cells.

PPNDS is an agonist, not an antagonist, for the ATP receptor of Paramecium.

Paramecium represents a simple, eukaryotic model system to study the cellular effects of some neuroactive drugs. They respond to the agonist beta,gamma-methylene ATP with a transient depolarizing receptor potential, Ca(2+)-based action potentials and repetitive bouts of forward and backward swimming called 'avoiding reactions' (AR). In vivo [(32)P]ATP binding assays showed saturable [(32)P]ATP binding with an apparent K(d) of approximately 23 nmol l(-1). Prolonged (15 min) exposure to 25 micro mol l(-1) beta,gamma-methylene ATP caused behavioral adaptation and losses of AR, ATP receptor potentials and [(32)P]ATP binding. While screening various ATP receptor inhibitors, we found that the P2X1 'antagonist' pyridoxal-phosphate naphthylazo-nitro-disulfate (PPNDS) is actually an agonist, producing the same responses as beta,gamma-methylene ATP. [(32)P]ATP binding assays suggest that both agonists may bind to the same site as [(32)P]ATP. Cross-adaptation is also seen between PPNDS and beta,gamma-methylene ATP in terms of losses in AR, depolarizing receptor potentials and [(32)P]ATP binding. We conclude that the inhibition caused by PPNDS in Paramecium is due to agonist-induced desensitization. Either this represents a unique new class of ATP receptors, in which PPNDS is an agonist instead of an antagonist, or PPNDS (and other drugs like it) may actually be an agonist in many other cell types in which prolonged exposure is necessary for inhibition.