Plasmodium falciparum Erythrocytic Stage Parasites Require the Putative Autophagy Protein PfAtg7 for Normal Growth

Analysis of the Plasmodium falciparum genome reveals a limited number of putative autophagy genes, specifically the four genes involved in ATG8 lipidation, an essential step in formation of autophagosomes. In yeast, Atg8 lipidation requires the E1-type ligase Atg7, an E2-type ligase Atg3, and a cysteine protease Atg4. These four putative P. falciparum ATG (PfATG) genes are transcribed during the parasite’s erythrocytic stages. PfAtg7 has relatively low identity and similarity to yeast Atg7 (14.7% and 32.2%, respectively), due primarily to long insertions typical of P. falciparum. Excluding the insertions the identity and similarity are higher (38.0% and 70.8%, respectively). This and the fact that key residues are conserved, including the catalytic cysteine and ATP binding domain, we hypothesize that PfAtg7 is the activating enzyme of PfAtg8. To assess the role of PfAtg7 we have generated two transgenic parasite lines. In one, the PfATG7 locus was modified to introduce a C-terminal hemagglutinin tag. Western blotting reveals two distinct protein species, one migrating near the predicted 150 kDa and one at approximately 65 kDa. The second transgenic line introduces an inducible degradation domain into the PfATG7 locus, allowing us to rapidly attenuate PfAtg7 protein levels. Corresponding species are also observed in this parasite line at approximately 200 kDa and 100 kDa. Upon PfATG7 attenuation parasites exhibit a slow growth phenotype indicating the essentiality of this putative enzyme for normal growth.     

Correction to: External Supplement of Impulsive Micromanager Trichoderma Helps in Combating CO2 Stress in Rice Grown Under FACE

Plant Molecular Biology Reporter - The original version of this article unfortunately contained missing information at author’s affiliations. The affiliation address of the author’s...     

Diverging functions of Scr between embryonic and post-embryonic development in a hemimetabolous insect, Oncopeltus fasciatus

Hemimetabolous insects undergo an ancestral mode of development in which embryos hatch into first nymphs that resemble miniature adults. While recent studies have shown that homeotic (hox) genes establish segmental identity of first nymphs during embryogenesis, no information exists on the function of these genes during post-embryogenesis. To determine whether and to what degree hox genes influence the formation of adult morphologies, we performed a functional analysis of Sex combs reduced (Scr) during post-embryonic development in Oncopeltus fasciatus. The main effect was observed in prothorax of Scr-RNAi adults, and ranged from significant alterations in its size and shape to a near complete transformation of its posterior half toward a T2-like identity. Furthermore, while the consecutive application of Scr-RNAi at both of the final two post-embryonic stages (fourth and fifth) did result in formation of ectopic wings on T1, the individual applications at each of these stages did not. These experiments provide two new insights into evolution of wings. First, the role of Scr in wing repression appears to be conserved in both holo- and hemimetabolous insects. Second, the prolonged Scr-depletion (spanning at least two nymphal stages) is both necessary and sufficient to restart wing program. At the same time, other structures that were previously established during embryogenesis are either unaffected (T1 legs) or display only minor changes (labium) in adults. These observations reveal a temporal and spatial divergence of Scr roles during embryonic (main effect in labium) and post-embryonic (main effect in prothorax) development.     

A Well-Defined Readily Releasable Pool with Fixed Capacity for Storing Vesicles at Calyx of Held

The readily releasable pool (RRP) of vesicles is a core concept in studies of presynaptic function. However, operating principles lack consensus definition and the utility for quantitative analysis has been questioned. Here we confirm that RRPs at calyces of Held from 14 to 21 day old mice have a fixed capacity for storing vesicles that is not modulated by Ca²⁺. Discrepancies with previous studies are explained by a dynamic flow-through pool, established during heavy use, containing vesicles that are released with low probability despite being immediately releasable. Quantitative analysis ruled out a posteriori explanations for the vesicles with low release probability, such as Ca²⁺-channel inactivation, and established unexpected boundary conditions for remaining alternatives. Vesicles in the flow-through pool could be incompletely primed, in which case the full sequence of priming steps downstream of recruitment to the RRP would have an average unitary rate of at least 9/s during heavy use. Alternatively, vesicles with low and high release probability could be recruited to distinct types of release sites; in this case the timing of recruitment would be similar at the two types, and the downstream transition from recruited to fully primed would be much faster. In either case, further analysis showed that activity accelerates the upstream step where vesicles are initially recruited to the RRP. Overall, our results show that the RRP can be well defined in the mathematical sense, and support the concept that the defining mechanism is a stable group of autonomous release sites.     

Microsatellite repeat dynamics in mitochondrial genomes of phytopathogenic fungi: frequency and distribution in the genic and intergenic regions

The frequency and distribution of microsatellites were analyzed in the 19 mitogenomes of phytopathogenic fungi covering five phyla. Our analysis revealed that in all the mitogenomes studied, the frequency and relative abundance varied, and it was neither influenced by genome size nor by GC content. SSRs were found to be differential distributed in genic and intergenic regions. An average of 5.14 (23.6%) SSRs were present in genic sequences and 21.7 (76.4%) SSRs were located in the intergenic sequences. Relative abundance of SSRs in mitogenomes was the highest in Aspergillus tubigensis, whereas, it was the least in Phaeosphaeria nodurum, the average being 0.45. Trinucleotide repeats were the most abundant motifs in the genic and intergenic regions of the mitogenomes of the phytopathogenic fungi. Among the genes, cox1 harbors the maximum SSRs, whereas cox3 and nad 7 contain the least. Based on the presence of SSRs in a particular gene, genetic relationships among individual organisms were also established.     

Salinity–induced modulations in the protective defense system and programmed cell death in <Emphasis Type="Italic">Nostoc muscorum</Emphasis>

To study the biochemical adaptive responses of the blue green algae Nostoc muscorum to the salinity- induced stress they were exposed to various concentrations (5, 10, 15, 20 or 200 mM) of sodium chloride (NaCl). A dose-dependent inhibition of total protein content showed an adverse effect of NaCl on the growth of N. muscorum. Four-day treatment of NaCl (5–20 mM) progressively increased the content of the total peroxide with subsequent increase of the superoxide dismutase (SOD) activity, proline and total phenol content only up to 10 mM NaCl. Higher concentrations of NaCl caused significant decrease in both the enzymatic and non-enzymatic antioxidants. Induction of two polypeptides of ~29.10 and 40.15 kD as well as upregulation of many polypeptides as compared to control indicates the induction of SOD and dehydrin-like proteins, which supports the theory of adaptation against the salt stress. Furthermore, adaptation of N. muscorum to lower concentrations (5–20 mM) of NaCl was also confirmed by no fragmentation of DNA while DNA fragmentation indicating programmed cell death (PCD) could only be seen at 200 mM NaCl for 12 hours. We hypothesized that proline may confer a positive role to combat salinity stress and the same was confirmed by treatment of the test blue green algae with exogenous proline (1 and 10 μM). The results exhibited 16% reduction in the level of total peroxides, which is a well known oxidative stress marker in the 10 μM proline-treated NaCl group as compared to direct exposure to NaCl.     

A comparative study of microsatellites among crocodiles and development of genomic resources for the critically endangered Indian gharial

Genetica - Next-generation sequencing has allowed us to explore new methods, where comparative and population genomics can be used simultaneously. Keeping this in mind, we surveyed and analyzed the...     

RNAi analysis of nubbin embryonic functions in a hemimetabolous insect,Oncopeltus fasciatus

SUMMARY Although the expression of the POU homeodomain gene nubbin (nub) has been examined in several arthropod species, its function has been studied only in Drosophila. Here, we provide the first insight into functional roles of this gene in a hemimetabolous insect species, Oncopeltus fasciatus. The analysis of its function using RNAi resulted in the altered morphology of antennae and labial tubes in the head, legs in the thorax, and, most notably, the growth of ectopic appendages originating from abdominal segments A2–A6. This change in the morphology of the abdomen can largely be attributed to the altered expression patterns of two hox genes, Ubx and abd‐A, in RNAinub embryos. First, abd‐A expression is completely abolished in A3–A6. Second, weak Ubx expression expands posteriorly to encompass novel domains in A2 and A3. Concomitant with these changes, limbs on A2 and A3 are small and less developed, whereas limbs on A4–A6 are large thoracic‐like legs. These results show that nub function is necessary for normal abd‐A expression and thus plays a critical role in suppressing leg formation on the abdomen. The loss of this regulation leads to upregulation of Distal‐less, and subsequent development of appendages. In Drosophila, however, abd‐A expression is unaffected in a nub‐depleted background, indicating that no such regulatory relationship exists between these two genes in the fruit fly. These differences reveal that variation exists in the genetic mechanisms that maintain an ancient insect feature, the limbless abdomen.