Aishe A. Sarshad

University of Gothenburg | GU · Department of Medical Biochemistry and Cell Biology

In mammals, RNA interference (RNAi) is mostly studied as a cytoplasmic event, however, numerous reports convincingly show nuclear localization of the AGO proteins. Nevertheless, the mechanism of nuclear entry remains to be fully elucidated, and the extent of nuclear RNAi further explored. We found that reduced Lamin A levels significantly induced n...

Argonaute proteins (AGOs) play crucial roles in RNA-induced silencing complex (RISC) formation and activity. AGOs loaded with small RNA molecules (miRNA or siRNA) either catalyze endoribonucleolytic cleavage of target RNAs or recruit factors responsible for translational silencing and target destabilization. miRNAs are well characterized and broadl...

Born to be a wildling Inbred laboratory mouse strains are used extensively in basic and translational immunology research. However, the commensal and pathogenic repertoire of resident microbes encountered in the wild is not replicated in a lab setting. This can substantially distort how the immune system develops and functions, leading to false ass...

We have encountered azygiid cercariae shed from the snail, Pleurocera semicarinata, at North Elkhorn Creek, Scott County, Kentucky, since 2004. Samples of these cercariae were frozen, and their DNA analyzed. The resulting alignment total length of 394 base pairs from partial 5.8S and 28S genes and the complete ribosomal internal transcribed spacer...

Dysregulation of miRNA expression is associated with multiple diseases, including cancers, in which small RNAs can have either oncogenic or tumor suppressive functions. Here we investigated the potential tumor suppressive function of miR-450a, one of the most significantly downregulated miRNAs in ovarian cancer. RNA-seq analysis of the ovarian canc...

Many small-interfering (si)RNAs are toxic to cancer cells through a 6mer seed sequence (positions 2–7 of the guide strand). Here we performed an siRNA screen with all 4096 6mer seeds revealing a preference for guanine in positions 1 and 2 and a high overall G or C content in the seed of the most toxic siRNAs for four tested human and mouse cell lin...

CD95/Fas ligand binds to the death receptor CD95 to induce apoptosis in sensitive cells. We previously reported that CD95L mRNA is enriched in sequences that, when converted to si/shRNAs, kill all cancer cells by targeting critical survival genes (Putzbach et al., 2017). We now report expression of full-length CD95L mRNA itself is highly toxic to c...

In mammals, gene silencing by the RNA-induced silencing complex (RISC) is a well-understood cytoplasmic posttranscriptional gene regulatory mechanism. Here, we show that embryonic stem cells (ESCs) contain high levels of nuclear AGO proteins and that in ESCs nuclear AGO protein activity allows for the onset of differentiation. In the nucleus, AGO p...

The enhancer regions of the myogenic master regulator MyoD give rise to at least two enhancer RNAs. Core enhancer eRNA (CEeRNA) regulates transcription of the adjacent MyoD gene, whereas DRReRNA affects expression of Myogenin in trans. We found that DRReRNA is recruited at the Myogenin locus, where it colocalizes with Myogenin nascent transcripts....

Many siRNAs and shRNAs induce a form of cell death in all cancer cells that involves silencing a set of critical survival genes, a process called death induced by survival gene elimination (DISE). Mechanistically a 6mer seed sequence (position 2-7 of the guide strand) is sufficient to confer DISE-inducing activity. We have now performed a strand sp...

CD95/Fas ligand (CD95L) is best characterized for its role in activating extrinsic apoptosis through binding to its receptor, CD95. However, we recently reported that the CD95L mRNA is enriched in sequences that when converted to si- or shRNAs kill cancer cells through an RNAi-dependent mechanism. When loaded into the RNA Induced Silencing Complex...

CD95/Fas ligand binds to the death receptor CD95/Fas to induce apoptosis in sensitive cells. We previously reported the CD95L mRNA is enriched in sequences that, when converted to si/shRNAs, are toxic to cells (Putzbach et al., 2017). These si/shRNAs kill all cancer cells through a RNAi off-target effect by targeting critical survival genes. We now...

Results of the RNA-Seq analysis used to generate Figure 4B.

The 6mer and 8mer toxicity index.

Correlation between experimental shRNA toxicity and TI.

Gene lists used in this work.

Over 80% of multiple-tested siRNAs and shRNAs targeting CD95 or CD95 ligand (CD95L) induce a form of cell death characterized by simultaneous activation of multiple cell death pathways preferentially killing transformed and cancer stem cells. We now show these si/shRNAs kill cancer cells through canonical RNAi by targeting the 3’UTR of critical sur...

Over 80% of multiple tested siRNAs and shRNAs targeting CD95 or CD95 ligand (CD95L) induce a form of cell death characterized by simultaneous activation of multiple cell death pathways preferentially killing transformed and cancer stem cells. We now show these si/shRNAs kill cancer cells through canonical RNAi by targeting the 3’UTR of critical sur...

Ribosome biogenesis is essential for cell growth and proliferation and is commonly elevated in cancer. Accordingly, numerous oncogene and tumor suppressor signaling pathways target rRNA synthesis. In breast cancer, non-canonical Wnt signaling by Wnt5a has been reported to antagonize tumor growth. Here, we show that Wnt5a rapidly represses rDNA gene...

The study of protein-RNA interactions is critical for our understanding of cellular processes and regulatory circuits controlled by RNA binding proteins (RBPs). Recent next generation sequencing-based approaches significantly promoted our understanding of RNA biology and its importance for cell function. We present a streamlined protocol for Photoa...

Author Synthesis of the translation machinery, including the mega-Dalton, RNA-protein ribosome complex, serves as a key driver of cellular growth and proliferation. It is therefore unsurprising that ribosomal biogenesis is under intricate regulation. The process through which ribosomes are made entails the coordination of components from diverse s...

Wnt5a decreases nucleoli size via non-canonical Wnt signaling and reduces cell growth. (a) Immunofluorescence analysis of β-catenin (green) staining after treatment of BT549 cells for 3 hours with vehicle, Wnt3a (200 ng/mL) or Wnt5a (200 ng/mL). The inability of Wnt5a treatment to induce stabilization and nuclear translocation of β-catenin (green),...

Nucleolar localization of DVL1. (a) Immunofluorescence with rabbit polyclonal antibody for DVL1 (red) merged with DNA (blue) in MCF7 and MDA-MB-231 breast cancer cells. Scale bar, 10 μm. (n = 3). (b) Exogenous DVL1 ectopically expressed in Rat2 cells localizes to the nucleolus. Immunofluorescence of DVL1 (green) and Fibrillarin (red) and their co-l...

DVL2 and DVL3 are excluded from nucleoli. (a) Immunofluorescence and confocal microscopy using antibodies to DVL1 (green) and Fibrillarin (red) merged with DNA (blue) in MCF7 cells and MCF7 cells stably expressing Wnt5a treated with ActD at 1000 ng/mL for 4 hours. (b) Immunofluorescence and confocal microscopy using antibodies to DVL2 (red) and UBF...

Knockdown of endogenous DVL1 does not affect DVL2 or DVL3 levels. (a) Western blots showing specific reduction of DVL1 protein levels, but no change in DVL2 or DVL3, in BT549 and MCF7 cells transduced with DVL1 shRNA (shDVL1) compared to non-silencing shRNA (Ctrl). Tubulin and GAPDH served as loading controls (n = 3). (b) Nucleofection of MCF7 cell...

RNA-protein interactions are central to cellular homeostasis and control every aspect of RNA metabolism in the cell. A variety of in vitro and in vivo techniques have been developed in the last three decades to study these interactions. Here we provide a brief review of the currently available techniques as well as an in-depth discussion of experim...

Actin and nuclear myosin 1 (NM1) are regulators of transcription and chromatin organization. Using a genome-wide approach, we report here that β-actin binds intergenic and genic regions across the mammalian genome, associated with both protein-coding and rRNA genes. Within the rDNA, the distribution of β-actin correlated with NM1 and the other subu...

Nuclear myosin 1c (NM1) is emerging as regulator of transcription and chromatin organization. Using chromatin immunoprecipitation and deep sequencing (ChIP-Seq) in combination with molecular analyses, here we investigated the global association of NM1 with the mammalian genome. Analysis of the ChIP-Seq data demonstrates that NM1 binds across the en...

In the eukaryotic cell nucleus, actin and myosin are emerging as essential regulators of nuclear function. At gene level, they regulate chromatin and modulate RNA polymerase transcription, and at the RNA level, they are involved in the metabolism of ribonucleoprotein complexes. Furthermore, actin and myosin are involved in maintaining the structure...

Author Summary Nuclear actin and myosin are essential regulators of gene expression. At the exit of mitosis, nuclear myosin 1c (NM1) mediates RNA polymerase I (pol I) transcription activation and cell cycle progression by modulating assembly of the chromatin remodeling complex WICH with the subunits WSTF and SNF2h and, crucially, facilitating H3K9...

Author Summary Actin and myosin are key regulators of several processes that occur in the cell nucleus. In rRNA biogenesis, actin in complex with nuclear myosin 1c (NM1) is involved in several phases of rDNA transcription. Further, NM1 interacts with the chromatin remodelling complex WICH, with the subunits WSTF and SNF2h. The multiprotein assembly...

The objectives of this study were to describe the effect of light wavelength and osmolality on the vertical swimming distance of the cercaria of Proterometra macrostoma. Significant differences were found in the average swimming distances and electrical burst activity in the tail of cercariae exposed to red vs. blue, green or white light, but no di...

The objectives of this study were to describe the (1) location of Proterometra macrostoma rediae in the snail intermediate host, Elimia semicarinata, and (2) daily emergence of cercariae from individual snails over 3 weeks. Decalcification and histological sections of infected snails revealed rediae restricted to the bottom whorl within the mantle...

The objectives of the present study were (1) to compare the frequency of Proterometra macrostoma (Faust) strains and the number of single vs. multiple strain infections in snails, Elimia semicarinata, collected from North Elkhorn Creek during June 2000, 2004, and 2007, (2) to measure selected cercarial strain egg loads and swimming distance under r...