Bidirectional silencing of RNA polymerase I transcription by a strand switch region in Trypanosoma brucei

The procyclin genes in Trypanosoma brucei are transcribed by RNA polymerase I as part of 5-10 kb long polycistronic transcription units on chromosomes VI and X. Each procyclin locus begins with two procyclin genes followed by at least one procyclin-associated gene (PAG). In procyclic (insect midgut) form trypanosomes, PAG mRNA levels are about 100-fold lower than those of procyclins. We show that deletion of PAG1, PAG2 or PAG3 results in increased mRNA levels from downstream genes in the same transcription unit. Nascent RNA analysis revealed that most of the effects are due to increased transcription elongation in the knockouts. Furthermore, transient and stable transfections showed that sequence elements on both strands of PAG1 can inhibit Pol I transcription. Finally, by database mining we identified 30 additional PAG-related sequences that are located almost exclusively at strand switch regions and/or at sites where a change of RNA polymerase type is likely to occu

Spillovers of Prosocial Motivation: Evidence from an Intervention Study on Blood Donors

Blood donations are increasingly important for medical procedures, while meeting demand is challenging. This paper studies the role of spillovers arising from social interactions in the context of voluntary blood donations. We analyze a large scale intervention among pairs of blood donors who live at the same street address. A quasi-random phone call provides the instrument for identifying the extent to which the propensity to donate spills over within these pairs. Spillovers transmit 41% to 46% of the behavioral impulse from one donor to the peer. This creates a significant social multiplier, ranging between 1.7 and 1.85. There is no evidence that these spillovers lead to intertemporal substitution. Taken together, our findings indicate that policy interventions have a substantially larger effect when targeted towards pairs instead of isolated individuals

Analysis of C. elegans intestinal gene expression and polyadenylation by fluorescence-activated nuclei sorting and 3′-end-seq

Despite the many advantages of Caenorhabditis elegans, biochemical approaches to study tissue-specific gene expression in post-embryonic stages are challenging. Here, we report a novel experimental approach for efficient determination of tissue-specific transcriptomes involving the rapid release and purification of nuclei from major tissues of post-embryonic animals by fluorescence-activated nuclei sorting (FANS), followed by deep sequencing of linearly amplified 3′-end regions of transcripts (3′-end-seq). We employed these approaches to compile the transcriptome of the developed C. elegans intestine and used this to analyse tissue-specific cleavage and polyadenylation. In agreement with intestinal-specific gene expression, highly expressed genes have enriched GATA-elements in their promoter regions and their functional properties are associated with processes that are characteristic for the intestine. We systematically mapped pre-mRNA cleavage and polyadenylation sites, or polyA sites, including more than 3000 sites that have previously not been identified. The detailed analysis of the 3′-ends of the nuclear mRNA revealed widespread alternative polyA site use (APA) in intestinally expressed genes. Importantly, we found that intestinal polyA sites that undergo APA tend to have U-rich and/or A-rich upstream auxiliary elements that may contribute to the regulation of 3′-end formation in the intestin

Bidirectional silencing of RNA polymerase I transcription by a strand switch region in Trypanosoma brucei

The procyclin genes in Trypanosoma brucei are transcribed by RNA polymerase I as part of 5–10 kb long polycistronic transcription units on chromosomes VI and X. Each procyclin locus begins with two procyclin genes followed by at least one procyclin-associated gene (PAG). In procyclic (insect midgut) form trypanosomes, PAG mRNA levels are about 100-fold lower than those of procyclins. We show that deletion of PAG1, PAG2 or PAG3 results in increased mRNA levels from downstream genes in the same transcription unit. Nascent RNA analysis revealed that most of the effects are due to increased transcription elongation in the knockouts. Furthermore, transient and stable transfections showed that sequence elements on both strands of PAG1 can inhibit Pol I transcription. Finally, by database mining we identified 30 additional PAG-related sequences that are located almost exclusively at strand switch regions and/or at sites where a change of RNA polymerase type is likely to occur

The Sting of Rejection: Deferring Blood Donors due to Low Hemoglobin Values Reduces Future Returns

Background: Roughly one quarter of short-term temporary deferrals (STTD) of blood donors are low-hemoglobin deferrals (LHD), i.e. STTD due to a hemoglobin (Hb) value falling below a cutoff of 125 g/L for female and 135 g/L for male donors. Since voluntarily donating blood is a prosocial activity, donors may perceive deferral as social exclusion, which can cause social pain, decrease self-esteem, and lead to antisocial behavior. However, little is known about the causal impacts of LHD on donor return. Study Design and Methods: We conducted a quasi-experiment with 80,060 donors invited to blood drives in the canton of Zurich, Switzerland, between 2009 and 2014. Within a narrow window of Hb values around the predetermined cutoff, the rate of LHD jumps discontinuously. This discontinuous jump allows us to quantify the causal effects of LHD on donor return, as it is uncorrelated with other unobserved factors that may also affect donor return. Results: We found different behavioral reactions to LHD for female and male donors. Female donors do not react to the first LHD. However, after any repeated LHD, they are 13.53 percentage points (p <0.001) less likely to make at least 1 donation attempt within the next 18 months and make 0.389 fewer donation attempts (p <0.001). Male donors react to the first LHD. They are 5.32 percentage points (p = 0.139) less likely to make at least 1 donation attempt over the next 18 months and make 0.227 (p = 0.018) fewer donation attempts. After any repeated LHD, male donors are 13.30 percentage points (p = 0.004) less likely to make at least 1 donation attempt and make 0.152 (p = 0.308) fewer donation attempts. Conclusion: LHD have detrimental impacts on donor return, especially if they occur repeatedly – suggesting that avoiding false LHD and helping donors to better cope with them helps to maintain the pool of prospective donors

Regulation of transcription termination in the nematode Caenorhabditis elegans

The current predicted mechanisms that describe RNA polymerase II (pol II) transcription termination downstream of protein expressing genes fail to adequately explain, how premature termination is prevented in eukaryotes that possess operon-like structures. Here we address this issue by analysing transcription termination at the end of single protein expressing genes and genes located within operons in the nematode Caenorhabditis elegans. By using a combination of RT-PCR and ChIP analysis we found that pol II generally transcribes up to 1 kb past the poly(A) sites into the 3′ flanking regions of the nematode genes before it terminates. We also show that pol II does not terminate after transcription of internal poly(A) sites in operons. We provide experimental evidence that five randomly chosen C. elegans operons are transcribed as polycistronic pre-mRNAs. Furthermore, we show that cis-splicing of the first intron located in downstream positioned genes in these polycistronic pre-mRNAs is critical for their expression and may play a role in preventing premature pol II transcription termination

Molecular mechanism of poly(ADP-ribosyl)ation by PARP1 and identification of lysine residues as ADP-ribose acceptor sites

Poly(ADP-ribose) polymerase 1 (PARP1) synthesizes poly(ADP-ribose) (PAR) using nicotinamide adenine dinucleotide (NAD) as a substrate. Despite intensive research on the cellular functions of PARP1, the molecular mechanism of PAR formation has not been comprehensively understood. In this study, we elucidate the molecular mechanisms of poly(ADP-ribosyl)ation and identify PAR acceptor sites. Generation of different chimera proteins revealed that the amino-terminal domains of PARP1, PARP2 and PARP3 cooperate tightly with their corresponding catalytic domains. The DNA-dependent interaction between the amino-terminal DNA-binding domain and the catalytic domain of PARP1 increased Vmax and decreased the Km for NAD. Furthermore, we show that glutamic acid residues in the auto-modification domain of PARP1 are not required for PAR formation. Instead, we identify individual lysine residues as acceptor sites for ADP-ribosylation. Together, our findings provide novel mechanistic insights into PAR synthesis with significant relevance for the different biological functions of PARP family members

PARP1 ADP-ribosylates lysine residues of the core histone tails

The chromatin-associated enzyme PARP1 has previously been suggested to ADP-ribosylate histones, but the specific ADP-ribose acceptor sites have remained enigmatic. Here, we show that PARP1 covalently ADP-ribosylates the amino-terminal histone tails of all core histones. Using biochemical tools and novel electron transfer dissociation mass spectrometric protocols, we identify for the first time K13 of H2A, K30 of H2B, K27 and K37 of H3, as well as K16 of H4 as ADP-ribose acceptor sites. Multiple explicit water molecular dynamics simulations of the H4 tail peptide into the catalytic cleft of PARP1 indicate that two stable intermolecular salt bridges hold the peptide in an orientation that allows K16 ADP-ribosylation. Consistent with a functional cross-talk between ADP-ribosylation and other histone tail modifications, acetylation of H4K16 inhibits ADP-ribosylation by PARP1. Taken together, our computational and experimental results provide strong evidence that PARP1 modifies important regulatory lysines of the core histone tails

Light Harvesting Schemes for High Efficiency Thin Film Silicon Solar Cells

In Thin Film Silicon (TF-Si) solar cells light harvesting schemes must guarantee an efficient light trapping in the thin absorber layers without decreasing the silicon layers quality and consecutively the p-i-n diodes electrical performance. TF-Si solar cells resilience to the substrate roughness is reported to be possibly improved through optimizations of the cell design and of the silicon deposition processes. By further tailoring the superstrate texture, amorphous silicon / microcrystalline silicon (a-Si:H/mu c-Si:H) tandem solar cells with an initial efficiency up to 13.7 % and a stabilized efficiency up to 11.8 % are demonstrated on single-scale textured superstrates. An alternative approach combining large and smooth features nanoimprinted onto a transparent lacquer with small and sharp textures from as-grown LPCVD ZnO is then shown to have a high potential for further increasing TF-Si devices efficiency. First results demonstrate up to 14.1 % initial efficiency for a TF-Si tandem solar cell

Major Surface Glycoproteins of Insect Forms of Trypanosoma brucei Are Not Essential for Cyclical Transmission by Tsetse

Procyclic forms of Trypanosoma brucei reside in the midgut of tsetse flies where they are covered by several million copies of glycosylphosphatidylinositol-anchored proteins known as procyclins. It has been proposed that procyclins protect parasites against proteases and/or participate in tropism, directing them from the midgut to the salivary glands. There are four different procyclin genes, each subject to elaborate levels of regulation. To determine if procyclins are essential for survival and transmission of T. brucei, all four genes were deleted and parasite fitness was compared in vitro and in vivo. When co-cultured in vitro, the null mutant and wild type trypanosomes (tagged with cyan fluorescent protein) maintained a near-constant equilibrium. In contrast, when flies were infected with the same mixture, the null mutant was rapidly overgrown in the midgut, reflecting a reduction in fitness in vivo. Although the null mutant is patently defective in competition with procyclin-positive parasites, on its own it can complete the life cycle and generate infectious metacyclic forms. The procyclic form of T. brucei thus differs strikingly from the bloodstream form, which does not tolerate any perturbation of its variant surface glycoprotein coat, and from other parasites such as Plasmodium berghei, which requires the circumsporozoite protein for successful transmission to a new host