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Mycotoxines are widespread toxic substances produced by moulds in human and animal foodstuffs. Some of them, such as aflatoxin B1 (AFB1), are very toxic even in small amounts, but the modern approach to the problem (and the link with public health) started only in the 60s. The presence of aflatoxin M1 (AFM1), in milk and dairy products is also a problem of crucial interest. AFM1 is resistant to thermal and chemical treatments, so mineral sorbents are highly advisable. Clay minerals are suitable, but only a few data are available, as most studies deal with AFB1 (absent in milk). The capacity of clay materials to remove or attenuate the contamination of AFM1 in bovine milk was studied, also considering the effects of these clays on the nutritional properties of milk (in terms of protein, fat and lactose). In order to make the experiments transferable for practical use, only clays available from the market were tested, as well a very simple clay-milk interaction procedure was used. Some bentonites and a kaolin were selected for the experimental work. In particular, different bentonites were selected in order to partially accomplish the wide crystal-chemical features of smectite. Kaolin was less effective than bentonites, but was still able to detoxify contaminated milk, even using a little amount of kaolin (2.4% of the milk suspension). The detoxification role of the bentonites used was variable, but still very efficient: contaminated bovine milk (up to about 80 ng/L) was purified to safe levels (50ng/L for adults and 25ng/L for lactants), with moderate alteration of the nutritional properties of milk; moreover, the bentonite residue in the purified milk was very low (0.4%). The protein was more sensitive to clay adsorption compared to fat and lactose, and the decrease of protein in the treated milk increased with the clay-milk ratio. Among bentonites (beidellite-montmorillonite, ferruginous and saponite types), a saponite clay showed the highest sorbent capacity, in agreement with theoretical consideration about its higher cell surface (available for AFM1) and higher hydrophobicity. Isothermal adsorption procedures for the two more effective bentonites were also carried out in water. The collected data show an effective and safe use of clays in the detoxification of milk (and dairy products) contaminated by AFM1. The climatic changes observed in the last decades point to a wider diffusion of mycotoxines and health effects, and push toward the search for safe, cheap and accessible food treatments.

Correct execution of mitosis in eukaryotes relies on timely activation and inactivation of cyclin B-dependent kinase 1 (cdk1), the M-phase-promoting factor (MPF). Once activated, MPF is sustained until mitotic spindle assembly by phosphorylation-dependent feedback loops that prevent inhibitory phosphorylation of cdk1 and ubiquitin-dependent degradation of cyclin B. Whether subsequent MPF inactivation and anaphase onset require a specific phosphatase(s) to reverse these feedback loops is not known. Here we show through biochemical and genetic evidence that timely MPF inactivation requires activity of the essential RNA polymerase II-carboxy-terminal domain phosphatase Fcp1, in a transcription-independent manner. We identify Cdc20, a coactivator of the ubiquitin ligase anaphase-promoting complex/cyclosome (APC/C) required for cyclin degradation and anaphase onset, USP44, a deubiquitinating peptidase that opposes APC/C action, and Wee1, a cdk1 inhibitory kinase, as relevant Fcp1 targets. We propose that Fcp1 has a crucial role in the liaison between dephosphorylation and ubiquitination that drives mitosis exit.

Anaphase initiation requires ubiquitin-dependent proteolysis of crucial substrates through activation of the ubiquitin ligase Anaphase promoting Complex/Cyclosome (ApC/C) in association with its coactivator Cdc20. To prevent chromosome segregation errors, effector proteins of a safeguard mechanism called spindle assembly checkpoint (SAC), Mad2 and BubR1, bind Cdc20 and restrain ApC/C(Cdc20) activation until spindle assembly. Coordinated chromosome segregation also requires timely SAC inactivation. Spindle assembly appears necessary to silence SAC, however, how resolution of the SAC effector branch is achieved is still largely unknown. We show here that the complex between Mad2 and Cdc20 peaked at prometaphase in mammalian cells, while its dissociation proceeded along with spindle assembly and required proteolysis. proteolysis did not appear required for assembly of metaphase spindles but rather needed for Mad2-Cdc20 complex resolution by promoting reversal of phosphorylations that maintain the complex. Indeed, in the absence of proteolysis, Mad2-Cdc20 complex dissociation was reversed by treatment with cyclin-dependent kinase or Aurora kinase inhibitors. Mad2-Cdc20 disassembly was, however, resistant to the potent pp1 and pp2A phosphatases inhibitor okadaic acid. We propose that SAC silencing in mammalian cells requires proteolysis-dependent activation of okadaic acid-resistant phosphatase(s) to reverse phosphorylations that lock the Mad2-Cdc20 complex.