Apicomplexan parasites are auxotrophic for a range of amino acids which must be salvaged from their host cells, either through direct uptake or degradation of host proteins. Here, we describe a family of plasma membrane-localized amino acid transporters, termed the Apicomplexan Amino acid Transporters (ApiATs), that are ubiquitous in apicomplexan parasites. Functional characterization of the ApiATs of Toxoplasma gondii indicate that several of these transporters are important for intracellular growth of the tachyzoite stage of the parasite, which is responsible for acute infections. We demonstrate that the ApiAT protein Tg ApiAT5-3 is an exchanger for aromatic and large neutral amino acids, with particular importance for L-tyrosine scavenging and amino acid homeostasis, and that Tg ApiAT5-3 is critical for parasite virulence. Our data indicate that T . gondii expresses additional proteins involved in the uptake of aromatic amino acids, and we present a model for the uptake and homeostasis of these amino acids. Our findings identify a family of amino acid transporters in apicomplexans, and highlight the importance of amino acid scavenging for the biology of this important phylum of intracellular parasites.

A long-established theoretical result states that, for a given total canopy nitrogen (N) content, canopy photosynthesis is maximized when the within-canopy gradient in leaf N per unit area (N(a)) is equal to the light gradient. However, it is widely observed that N(a) declines less rapidly than light in real plant canopies. Here we show that this general observation can be explained by optimal leaf acclimation to light subject to a lower-bound constraint on the leaf mass per area (m(a)). Using a simple model of the carbon-nitrogen (C-N) balance of trees with a steady-state canopy, we implement this constraint within the framework of the MAXX optimization hypothesis that maximizes net canopy C export. Virtually all canopy traits predicted by MAXX (leaf N gradient, leaf N concentration, leaf photosynthetic capacity, canopy N content, leaf-area index) are in close agreement with the values observed in a mature stand of Norway spruce trees (Picea abies L. Karst.). An alternative upper-bound constraint on leaf photosynthetic capacity (A(sat)) does not reproduce the canopy traits of this stand. MAXX subject to a lower bound on m(a) is also qualitatively consistent with co-variations in leaf N gradient, m(a) and A(sat) observed across a range of temperate and tropical tree species. Our study highlights the key role of constraints in optimization models of plant function.

Class I histone deacetylases (HDACs) participate in the regulation of DNA-templated processes such as transcription and replication. Members of this class can act locally at specific sites, or they can act more globally, contributing to a baseline acetylation state, both of which actions may be important for genome maintenance and organization. We previously identified a macronuclear-specific class I HDAC in Tetrahymena thermophila called Thd1p, which is expressed early in the development of the macronucleus when it initially becomes transcriptionally active. To test the idea that Thd1p is important for global chromatin integrity in an active macronucleus, Tetrahymena cells reduced in expression of Thd1p were generated. We observed phenotypes that indicated loss of chromatin integrity in the mutant cells, including DNA fragmentation and extrusion of chromatin from the macronucleus, variable macronuclear size and shape, enlarged nucleoli, and reduced phosphorylation of histone H1 from bulk chromatin. Macronuclei in mutant cells also contained more DNA. This observation suggests a role for Thd1p in the control of nuclear DNA content, a previously undescribed role for class I HDACs. Together, these phenotypes implicate Thd1p in the maintenance of macronuclear integrity in multiple ways, probably through site-specific changes in histone acetylation since no change in the acetylation levels of bulk histones was detected in mutant cells.

Class I histone deacetylases (HDACs) regulate DNA-templated processes such as transcription. They act both at specific loci and more generally across global chromatin, contributing to acetylation patterns that may underlie large-scale chromatin dynamics. Although hypoacetylation is correlated with highly condensed chromatin, little is known about the contribution of individual HDACs to chromatin condensation mechanisms. Using the ciliated protozoan Tetrahymena thermophila, we investigated the role of a specific class I HDAC, ⌻hd1p, in the reversible condensation of global chromatin. In this system, the normal physiological response to cell starvation includes the widespread condensation of the macronuclear chromatin and general repression of gene transcription. We show that the chromatin in Thd1p-deficient cells failed to condense during starvation. The condensation failure correlated with aberrant hyperphosphorylation of histone H1 and the overexpression of CDC2, encoding the major histone H1 kinase. Changes in the rate of acetate turnover on core histones and in the distribution of acetylated lysines 9 and 23/27 on histone H3 isoforms that were found to correlate with normal chromatin condensation were absent from Thd1p mutant cells. These results point to a role for a class I HDAC in the formation of reversible higher-order chromatin structures and global genome compaction through mechanisms involving the regulation of H1 phosphorylation and core histone acetylation/ deacetylation kinetics.

Apicomplexan parasites are auxotrophic for a range of amino acids which must be salvaged from their host cells, either through direct uptake or degradation of host proteins. Here, we describe a family of plasma membrane-localized amino acid transporters, termed the Apicomplexan Amino acid Transporters (ApiATs), that are ubiquitous in apicomplexan parasites. Functional characterization of the ApiATs of Toxoplasma gondii indicate that several of these transporters are important for intracellular growth of the tachyzoite stage of the parasite, which is responsible for acute infections. We demonstrate that the ApiAT protein TgApiAT5-3 is an exchanger for aromatic and large neutral amino acids, with particular importance for L-tyrosine scavenging and amino acid homeostasis, and that TgApiAT5-3 is critical for parasite virulence. Our data indicate that T. gondii expresses additional proteins involved in the uptake of aromatic amino acids, and we present a model for the uptake and homeostasis of these amino acids. Our findings identify a family of amino acid transporters in apicomplexans, and highlight the importance of amino acid scavenging for the biology of this important phylum of intracellular parasites.Author SummaryThe Apicomplexa comprise a large number of parasitic protozoa that have obligate intracellular lifestyles and cause significant human and animal diseases, including malaria, cryptosporidiosis, toxoplasmosis, coccidiosis in poultry, and various cattle fevers. Apicomplexans must scavenge essential nutrients from their hosts in order to proliferate and cause disease, including a range of amino acids. The direct uptake of these nutrients is presumed to be mediated by transporter proteins located in the plasma membrane of intracellular stages, although the identities of these proteins are poorly defined. Using a combination of bioinformatic, genetic, cell biological, and physiological approaches, we have characterized a family of plasma membrane-localized transporter proteins that we have called the Apicomplexan Amino acid Transporters (ApiATs). The family is found in apicomplexans and their closest free-living relatives. We show that TgApiAT5-3, a member of the family in the apicomplexan Toxoplasma gondii, is an exchanger for aromatic and large neutral amino acids. In particular, it is critical for uptake of tyrosine, and for parasite virulence in a mouse infection model. We conclude that ApiATs are a family of plasma membrane transporters that play crucial roles in amino acid scavenging by apicomplexan parasites.

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