Fire regime changes are considered a major threat to future biodiversity in the Mediterranean Basin. Such predictions remain untested, however, given that fire regime changes and their ecological impacts occur over timescales that are too long for direct observation. Here we analyse centennial-3 and millennial-scale shifts in fire regimes and compositional turnover to track the consequences of fire regime shifts on Mediterranean vegetation diversity. We estimated rate-of-change, richness and compositional turnover (beta diversity) in 13 selected high-resolution palaeoecological records from Mediterranean Iberia and compared these to charcoal-inferred fire regime changes. Event sequence analysis showed fire regime shifts to be significantly temporally associated with compositional turnover, particularly during the last three millennia. We find that the timing and direction of fire and diversity change in Mediterranean Iberia are best explained by long-term human-environment interactions dating back perhaps 7500 years. Evidence suggests that Neolithic burning propagated a first wave of increasing vegetation openness and promoted woodland diversity around early farming settlements. Landscape transformation intensified around 5500-5000 cal. yr BP and accelerated during the last two millennia, as fire led to permanent transitions in ecosystem state. These fire episodes increased open vegetation diversity, decreased woodland diversity and significantly altered richness on a regional scale. Our study suggests that anthropogenic fires played a primary role in diversity changes in Mediterranean Iberia. Their millennia-long legacy in today's vegetation should be considered for biodiversity conservation and landscape management.

Mutations were introduced in the motif 884 DDRW 887 from an extracellular peptide of the sodium pump ␣ subunit localized between M7 and M8 membrane spans to investigate a possible role of this structure in ion recognition. A homologous sequence 399 QDCW 402 that occurs in the P-loops of Na ؉ channels was shown earlier to be important for ion gating. Mutant sodium pumps were expressed in yeast and subsequently investigated for their behavior toward ouabain, Na ؉ , K ؉ , and ATP. Native enzyme and D884A, D884R, D885A, D885E, or D885R mutants all bind ouabain in the presence of phosphate and Mg 2؉ . The K D values determined from Scatchard analysis are in the range 5-8 nM for the native enzyme and the D884A, D885E, or D885A mutants, and 15.7 ؎ 2.04 and 30.1 ؎ 4.32 nM for mutants D884R and D885R, respectively. This ouabain binding is reduced in the presence of K ؉ in a similar way for both native or mutant sodium pumps with relative affinities (K 0.5 ) for K ؉ ranging from 1.4 to 3.7 mM. Ouabain binding in the presence of 100 M ATP is promoted by Na ؉ with K 0.5 ‫؍‬ 1.64 ؎ 0.01 mM for the native enzyme and K 0.5 ‫؍‬ 8.6 ؎ 1.35 mM for the D884R mutant. The K 0.5 values of the two enzymes for ATP are 0.66 ؎ 0.16 M and 1.1 ؎ 0.12 M, respectively. Ouabain binding as a function of Na ؉ concentration, on the other hand, is very low for the D885R mutant, even at an ATP concentration of 2 mM. Phosphate or eosin, however, are recognized by this mutant enzyme, so that a major conformational change within the ATP-binding site appears unlikely.The inability of the D885R mutant to bind ouabain in the presence of Na ؉ and ATP could be explained by assuming that the M7/M8 connecting extracellular loop, which also contains the mutated amino acids, is invaginated within the plane of the plasma membrane and possibly involved in acceptance and/or release of Na ؉ ions coming from cytosolic areas of the protein. In this case, the placement of an additional positive charge might repel Na ؉ ions and interrupt their flow, thus not allowing the enzyme to assume the proper conformational state for ouabain binding. Such invaginated hydrophilic protein structures, such as the P-loops of Na ؉ and K ؉ channels, are already known and have been shown to participate in ion conduction.The sodium pump from plasma membranes of animal cells is an oligomeric enzyme that consists of an ␣ and a ␤ subunit. The enzyme hydrolyzes ATP to actively transport Na ϩ ions out of the cell and K ϩ ions into the cell. Ouabain and other cardioactive steroids inhibit ATP hydrolysis and the transport of ions. The amino acids or the membrane spans involved in the formation of the ionophoric part of the sodium pump are not yet known although in several investigations mutations of single amino acids from the membrane spans of the ␣ subunit were shown to affect interactions of the enzyme with ions. Additional experiments, however, demonstrate that the ␤ subunit also affects the behavior of the enzyme toward the transported ions. Based on today's knowledge, interaction of ␣ a...

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At the turn of the second to the third millennium BC, fortified and systematically organized settlements along with a developed metallurgy emerged in the TransUral steppe. In order to reconstruct the related vegetation and climatic changes in the area and to detect effects of human impact during the respective Bronze-Age SintashtaPetrovka and Srubnaya-Alakul cultures (2100-1650 cal BC), palynological and sedimentological investigations accompanying archaeological excavations were carried out. Statistical analyses of pollen spectra from two sediment cores in the immediate vicinity of the settlement Kamennyi Ambar demonstrate substantial similarities in Bronze Age and present vegetation covers. Higher percentages of arboreal pollen, especially in respect of deciduous trees, and consistently high values of Artemisia within the regional spectra suggest that Bronze Age cultures developed in a relatively humid environment. Simultaneously, higher values of Chenopodiaceae, Plantago and Cichorioideae point to steppe degradation at local scales, whereas regional vegetation appears to have been relatively unaffected by human activities. Although more than 30 localities in the micro-region of Karagaily Ayat were analyzed, none of these sediment cores accounts for a continuous sedimentation during the last 9,000 years. Our results reveal that climatic fluctuations are not always directly reflected by sediment formation, with many nonclimatic factors, such as post-sedimentary processes, local morphology and erosion believed to be crucial. Only the use of high resolution AMS radiocarbon dating is suitable to detect hiatuses, hence providing a valuable clue to the interpretation of palaeoenvironmental conditions.