To characterize the regulatory pathways involved in the inhibition of cell differentiation induced by the impairment of mitochondrial activity, we investigated the relationships occurring between organelle activity and myogenesis using an avian myoblast cell line (QM7). The inhibition of mitochondrial translation by chloramphenicol led to a potent block of myoblast differentiation. Carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone and oligomycin, which affect the organelle at different levels, exerted a similar influence. In addition, we provided evidence that this phenomenon was not the result of an alteration in cell viability. Conversely, overexpression of the mitochondrial T3 receptor (p43) stimulated organelle activity and strongly potentiated myoblast differentiation. The involvement of mitochondrial activity in an actual regulation of myogenesis is further supported by results demonstrating that the muscle regulatory gene myogenin, in contrast to CMD1 (chicken MyoD) and myf5, is a specific transcriptional target of mitochondrial activity. Whereas myogenin mRNA and protein levels were down-regulated by chloramphenicol treatment, they were up-regulated by p43 overexpression, in a positive relationship with the expression level of the transgene. We also found that myogenin or CMD1 overexpression in chloramphenicol-treated myoblasts did not restore differentiation, thus indicating that an alteration in mitochondrial activity interferes with the ability of myogenic factors to induce terminal differentiation.Recent studies emphasize that mitochondria, in addition to their well known involvement in the regulation of energy metabolism, are implicated in the regulation of cell growth and differentiation. In particular, mitochondrial events are involved in the preliminary steps of apoptosis (1), and inhibition of mitochondrial activity, either by deleting mtDNA (rho°cells) or by blocking translation in the organelle, has been shown to stop or decrease the proliferation of different cell lines (2-4). Furthermore, the general activity of the organelle, not restricted to energy production, is implicated in such regulation (5, 6). In addition, mitochondrial protein synthesis inhibition is associated with the impairment of differentiation of different cell lines, such as mouse erythroleukemia (7) and mastocytoma cells (8), neurons (9), and human (10), avian (11) or murine myoblasts (12). In agreement with these data, several pathologies are associated with mitochondrial disorders, even if the links between mitochondrial genome rearrangements or activity and pathological symptoms are not always clearly established. Despite these reports, little is known about the molecular mechanisms involved in these regulations. First, the exclusive use of inhibitors of mitochondrial function in previous reports was not fully adapted to demonstrating the occurrence of an actual regulatory pathway involving mitochondrial activity in the regulation of cell differentiation. Second, the nature of the molecular signals underlying the...

In earlier research, we identified a 43-kDa c-ErbAalpha1 protein (p43) in the mitochondrial matrix of rat liver. In the present work, binding experiments indicate that p43 displays an affinity for triiodothyronine (T3) similar to that of the T3 nuclear receptor. Using in organello import experiments, we found that p43 is targeted to the organelle by an unusual process similar to that previously reported for MTF1, a yeast mitochondrial transcription factor. DNA-binding experiments demonstrated that p43 specifically binds to four mitochondrial DNA sequences with a high similarity to nuclear T3 response elements (mt-T3REs). Using in organello transcription experiments, we observed that p43 increases the levels of both precursor and mature mitochondrial transcripts and the ratio of mRNA to rRNA in a T3-dependent manner. These events lead to stimulation of mitochondrial protein synthesis. In transient-transfection assays with reporter genes driven by the mitochondrial D loop or two mt-T3REs located in the D loop, p43 stimulated reporter gene activity only in the presence of T3. All these effects were abolished by deletion of the DNA-binding domain of p43. Finally, p43 overexpression in QM7 cells increased the levels of mitochondrial mRNAs, thus indicating that the in organello influence of p43 was physiologically relevant. These data reveal a novel hormonal pathway functioning within the mitochondrion, involving a truncated form of a nuclear receptor acting as a potent mitochondrial T3-dependent transcription factor.

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Besides their involvement in the control of nuclear gene expression by activating several peroxisome proliferatoractivated receptors (PPARs), peroxisome proliferators influence mitochondrial activity. By analogy with the previous characterization of a mitochondrial T3 receptor (p43), we searched for the presence of a peroxisome proliferator target in the organelle. Using several antisera raised against different domains of PPARs, we demonstrated by Western blotting, immunoprecipitation and electron microscopy experiments, that a 45 kDa protein related to PPARQ Q2 (mt-PPAR) is located in the matrix of rat liver mitochondria. In addition, we found that the amounts of mt-PPAR are increased by clofibrate treatment. Moreover, in EMSA experiments mt-PPAR bound to a DR2 sequence located in the mitochondrial D-loop, by forming a complex with p43. Last, studies of tissue-specific expression indicated that mt-PPAR is detected in mitochondria of all tissues tested except the brain in amounts positively related to p43 abundance. ß

bAs in other crops, yield is an important trait to be selected for in edible mushrooms, but its inheritance is poorly understood. Therefore, we have investigated the complex genetic architecture of yield-related traits in Agaricus bisporus through the mapping of quantitative trait loci (QTL), using second-generation hybrid progeny derived from a cross between a wild strain and a commercial cultivar. Yield, average weight per mushroom, number of fruiting bodies per m 2 , earliness, and cap color were evaluated in two independent experiments. A total of 23 QTL were detected for 7 yield-related traits. These QTL together explained between 21% (two-flushes yield) and 59% (earliness) of the phenotypic variation. Fifteen QTL (65%) were consistent between the two experiments. Four regions underlying significant QTL controlling yield, average weight, and number were detected on linkage groups II, III, IV, and X, suggesting a pleiotropic effect or tight linkage. Up to six QTL were identified for earliness. The PPC1 locus, together with two additional genomic regions, explained up to 90% of the phenotypic variation of the cap color. Alleles from the wild parent showed beneficial effects for some yield traits, suggesting that the wild germ plasm is a valuable source of variation for several agronomic traits. Our results constitute a key step toward marker-assisted selection and provide a solid foundation to go further into the biological mechanisms controlling productive traits in the button mushroom.