Replication of the single-stranded DNA genome of plant geminiviruses follows a rolling circle mechanism. It strictly depends on a 'rolling circle replication initiator protein', the M(r) 41 kDa viral Rep protein, encoded by the C1 or AC1 genes. Using wheat dwarf virus (WDV) and tomato yellow leaf curl virus (TYLCV) as examples, we show that not only the full-size Rep proteins, but also a putative 30 kDa translation product of WDV open reading frame C1-N as well as an artificially shortened 24 kDa Rep of TYLCV, cleave and join single-stranded origin DNA in vitro. Thus the pivotal origin recognition and processing activities of geminivirus Rep proteins must be mediated by the amino-terminal domain of Rep.

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Dietary methionine restriction (MR) is well known to reduce body weight by increasing energy expenditure (EE) and insulin sensitivity. An elevated concentration of circulating fibroblast growth factor 21 (FGF21) has been implicated as a potential underlying mechanism. The aims of our study were to test whether dietary MR in the context of a high-fat regimen protects against type 2 diabetes in mice and to investigate whether vegan and vegetarian diets, which have naturally low methionine levels, modulate circulating FGF21 in humans. New Zealand obese (NZO) mice, a model for polygenic obesity and type 2 diabetes, were placed on isocaloric high-fat diets (protein, 16 kcal%; carbohydrate, 52 kcal%; fat, 32 kcal%) that provided methionine at control (Con; 0.86% methionine) or low levels (0.17%) for 9 wk. Markers of glucose homeostasis and insulin sensitivity were analyzed. Among humans, low methionine intake and circulating FGF21 levels were investigated by comparing a vegan and a vegetarian diet to an omnivore diet and evaluating the effect of a short-term vegetarian diet on FGF21 induction. In comparison with the Con group, MR led to elevated plasma FGF21 levels and prevented the onset of hyperglycemia in NZO mice. MR-fed mice exhibited increased insulin sensitivity, higher plasma adiponectin levels, increased EE, and up-regulated expression of thermogenic genes in subcutaneous white adipose tissue. Food intake and fat mass did not change. Plasma FGF21 levels were markedly higher in vegan humans compared with omnivores, and circulating FGF21 levels increased significantly in omnivores after 4 d on a vegetarian diet. These data suggest that MR induces FGF21 and protects NZO mice from high-fat diet–induced glucose intolerance and type 2 diabetes. The normoglycemic phenotype in vegans and vegetarians may be caused by induced FGF21. MR akin to vegan and vegetarian diets in humans may offer metabolic benefits via increased circulating levels of FGF21 and merits further investigation.—Castaño-Martinez, T., Schumacher, F., Schumacher, S., Kochlik, B., Weber, D., Grune, T., Biemann, R., McCann, A., Abraham, K., Weikert, C., Kleuser, B., Schürmann, A., Laeger, T. Methionine restriction prevents onset of type 2 diabetes in NZO mice.

The phage Mu transposase (MuA) binds to the ends of the Mu genome during the assembly of higher order nucleoprotein complexes. We investigate the structure and function of the MuA end-binding domain (Ibetagamma). The three-dimensional solution structure of the Ibeta subdomain (residues 77-174) has been determined using multidimensional NMR spectroscopy. It comprises five alpha-helices, including a helix-turn-helix (HTH) DNA-binding motif formed by helices 3 and 4, and can be subdivided into two interacting structural elements. The structure has an elongated disc-like appearance from which protrudes the recognition helix of the HTH motif. The topology of helices 2-4 is very similar to that of helices 1-3 of the previously determined solution structure of the MuA Igamma subdomain and to that of the homeodomain family of HTH DNA-binding proteins. We show that each of the two subdomains binds to one half of the 22 bp recognition sequence, Ibeta to the more conserved Mu end distal half (beta subsite) and Igamma to the Mu end proximal half (gamma subsite) of the consensus Mu end-binding site. The complete Ibetagamma domain binds the recognition sequence with a 100- to 1000-fold higher affinity than the two subdomains independently, indicating a cooperative effect. Our results show that the Mu end DNA-binding domain of MuA has a modular organization, with each module acting on a specific part of the 22 bp binding site. Based on the present binding data and the structures of the Ibeta and Igamma subdomains, a model for the interaction of the complete Ibetagamma domain with DNA is proposed.

Two tyrosines, both located in the amino-terminal domain of the protein, are conserved in geminivirus Rep proteins. The first one is part of the sequence motif-FLTY-whose function is still unknown, and the second one is the central amino acid of a sequence that has been suggested to catalyze replication initiation [33] (see Fig. 1) Here we provide biochemical proof that the tyrosine-103 of the Rep protein tomato yellow leaf curl virus (TYLCV) is the active amino acid of the cleavage/joining reaction.

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