Debra Black

Latent stem and progenitor cells in the hippocampus are activated by neural excitation.

The regulated production of neurons in the hippocampus throughout life underpins important brain functions such as learning and memory. Surprisingly, however, studies have so far failed to identify a resident hippocampal stem cell capable of providing the renewable source of these neurons. Here, we report that depolarizing levels of KCl produce a threefold increase in the number of neurospheres generated from the adult mouse hippocampus. Most interestingly, however, depolarizing levels of KCl led to the emergence of a small subpopulation of precursors (approximately eight per hippocampus) with the capacity to generate very large neurospheres (>250 μm in diameter). Many of these contained cells that displayed the cardinal properties of stem cells: multipotentiality and self-renewal. In contrast, the same conditions led to the opposite effect in the other main neurogenic region of the brain, the subventricular zone, in which neurosphere numbers decreased by ∼40% in response to depolarizing levels of KCl. Most importantly, we also show that the latent hippocampal progenitor population can be activated in vivo in response to prolonged neural activity found in status epilepticus. This work provides the first direct evidence of a latent precursor and stem cell population in the adult hippocampus, which is able to be activated by neural activity. Because the latent population is also demonstrated to reside in the aged animal, defining the precise mechanisms that underlie its activation may provide a means to combat the cognitive deficits associated with a decline in neurogenesis.

Co-Regulation of Cell Polarization and Migration by Caveolar Proteins PTRF/Cavin-1 and Caveolin-1

Caveolin-1 and caveolae are differentially polarized in migrating cells in various models, and caveolin-1 expression has been shown to quantitatively modulate cell migration. PTRF/cavin-1 is a cytoplasmic protein now established to be also necessary for caveola formation. Here we tested the effect of PTRF expression on cell migration. Using fluorescence imaging, quantitative proteomics, and cell migration assays we show that PTRF/cavin-1 modulates cellular polarization, and the subcellular localization of Rac1 and caveolin-1 in migrating cells as well as PKCα caveola recruitment. PTRF/cavin-1 quantitatively reduced cell migration, and induced mesenchymal epithelial reversion. Similar to caveolin-1, the polarization of PTRF/cavin-1 was dependent on the migration mode. By selectively manipulating PTRF/cavin-1 and caveolin-1 expression (and therefore caveola formation) in multiple cell systems, we unveil caveola-independent functions for both proteins in cell migration.

PTRF/cavin-1 neutralizes non-caveolar caveolin-1 microdomains in prostate cancer

Caveolin-1 has a complex role in prostate cancer and has been suggested to be a potential biomarker and therapeutic target. As mature caveolin-1 resides in caveolae, invaginated lipid raft domains at the plasma membrane, caveolae have been suggested as a tumor-promoting signaling platform in prostate cancer. However, caveola formation requires both caveolin-1 and cavin-1 (also known as PTRF; polymerase I and transcript release factor). Here, we examined the expression of cavin-1 in prostate epithelia and stroma using tissue microarray including normal, non-malignant and malignant prostate tissues. We found that caveolin-1 was induced without the presence of cavin-1 in advanced prostate carcinoma, an expression pattern mirrored in the PC-3 cell line. In contrast, normal prostate epithelia expressed neither caveolin-1 nor cavin-1, while prostate stroma highly expressed both caveolin-1 and cavin-1. Utilizing PC-3 cells as a suitable model for caveolin-1-positive advanced prostate cancer, we found that cavin-1 expression in PC-3 cells inhibits anchorage-independent growth, and reduces in vivo tumor growth and metastasis in an orthotopic prostate cancer xenograft mouse model. The expression of α-smooth muscle actin in stroma along with interleukin-6 (IL-6) in cancer cells was also decreased in tumors of mice bearing PC-3-cavin-1 tumor cells. To determine whether cavin-1 acts by neutralizing caveolin-1, we expressed cavin-1 in caveolin-1-negative prostate cancer LNCaP and 22Rv1 cells. Caveolin-1 but not cavin-1 expression increased anchorage-independent growth in LNCaP and 22Rv1 cells. Cavin-1 co-expression reversed caveolin-1 effects in caveolin-1-positive LNCaP cells. Taken together, these results suggest that caveolin-1 in advanced prostate cancer is present outside of caveolae, because of the lack of cavin-1 expression. Cavin-1 expression attenuates the effects of non-caveolar caveolin-1 microdomains partly via reduced IL-6 microenvironmental function. With circulating caveolin-1 as a potential biomarker for advanced prostate cancer, identification of the molecular pathways affected by cavin-1 could provide novel therapeutic targets.

Activation of latent precursors in the hippocampus is dependent on long-term potentiation

The recent discovery of a large latent population of precursor cells in the dentate gyrus of adult mice led us to investigate whether activation of this population is regulated by synaptic activity, thereby explaining the observation that environmental signals can affect neurogenesis. Using a variety of stimulation protocols, we found that only a long-term potentiation (LTP)-inducing protocol activated the latent precursor pool, leading to increased neurogenesis in the dentate gyrus. LTP induced by high-frequency stimulation (HFS) of the perforant pathway in vivo produced a two-fold increase in the number of neurospheres cultured from the stimulated hippocampus, compared with the unstimulated hippocampus. No increase in neurosphere number or neurogenesis was observed when the HFS failed to induce LTP. These results show that LTP can activate latent neural precursor cells in the adult mouse dentate gyrus, thereby providing a direct mechanism for regulating activity-driven neurogenesis. In the future, it may be possible to utilize such learning- or stimulation-induced neurogenesis to overcome disorders characterized by neuronal loss.

Isolation and characterisation of components of the Dunaliella tertiolecta chloroplast genome

Three chloroplast genes, psbA, psbB and rbcL, of the microalgae Dunaliella tertiolecta were targeted with the view to using these components in the construction of a chloroplast transformation vector. The three genes and surrounding genomic regions were isolated by screening libraries and using degenerate primers to amplify by PCR conserved coding regions and unknown flanking sequences. The putative Dunaliella psbA, psbB and rbcL proteins show high levels of sequence conservation sharing approximately 87, 92 and 97% similarity to the homologues of Chlamydomonas reinhardtii. Interestingly, four of the five introns of the psbA gene contain long open-reading frames which have sequence similarity to the H-N-H and GIY-YIG site-specific homing endonucleases suggesting that, like other microalgae, the Dunaliella gene contains group I introns. Putative promoter regions of the psbB and rbcL genes were isolated and found to contain the required signals necessary for gene expression.

Phenotypic and molecular dissection of Metaplastic Breast Cancer and the prognostic implications: Prognostic features of Metaplastic breast cancer.

Metaplastic breast carcinoma (MBC) is relatively rare but accounts for a significant proportion of global breast cancer mortality. This group is extremely heterogeneous and by definition exhibits metaplastic change to squamous and/or mesenchymal elements, including spindle, squamous, chondroid, osseous, and rhabdomyoid features. Clinically, patients are more likely to present with large primary tumours (higher stage), distant metastases, and overall, have shorter 5‐year survival compared to invasive carcinomas of no special type. The current World Health Organisation (WHO) diagnostic classification for this cancer type is based purely on morphology – the biological basis and clinical relevance of its seven sub‐categories are currently unclear. By establishing the Asia‐Pacific MBC (AP‐MBC) Consortium, we amassed a large series of MBCs (n = 347) and analysed the mutation profile of a subset, expression of 14 breast cancer biomarkers, and clinicopathological correlates, contextualising our findings within the WHO guidelines. The most significant indicators of poor prognosis were large tumour size (T3; p = 0.004), loss of cytokeratin expression (lack of staining with pan‐cytokeratin AE1/3 antibody; p = 0.007), EGFR overexpression (p = 0.01), and for ‘mixed’ MBC, the presence of more than three distinct morphological entities (p = 0.007). Conversely, fewer morphological components and EGFR negativity were favourable indicators. Exome sequencing of 30 cases confirmed enrichment of TP53 and PTEN mutations, and intriguingly, concurrent mutations of TP53, PTEN, and PIK3CA. Mutations in neurofibromatosis‐1 (NF1) were also overrepresented [16.7% MBCs compared to ∼5% of breast cancers overall; enrichment p = 0.028; mutation significance p = 0.006 (OncodriveFM)], consistent with published case reports implicating germline NF1 mutations in MBC risk. Taken together, we propose a practically minor but clinically significant modification to the guidelines: all WHO_1 mixed‐type tumours should have the number of morphologies present recorded, as a mechanism for refining prognosis, and that EGFR and pan‐cytokeratin expression are important prognostic markers. Copyright

Abstract 3040: Cavin-1 alters oncogenic effects of caveolin-1 microdomains in prostate cancer.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Caveolin-1 is associated with prostate cancer progression and has been suggested to be a biomarker and therapeutic target. Mature caveolin-1 resides in lipid raft domains at the plasma membrane, where it forms caveolae upon co-expression of cavin-1 (also known as PTRF; polymerase I and transcript release factor). In the absence of cavin-1, caveolin-1 does not form caveolae but are found on flat membrane. To determine if oncogenic caveolin-1 in prostate cancer is present in caveolae, we examined the relative expression of caveolin-1 and cavin-1 in normal, non-malignant and malignant prostate tissues. We found that caveolin-1 is induced in prostate cancer without cavin-1, an expression pattern mirror in the PC3 cell line. Previously we showed that expression of cavin-1 in PC3 cells recruits flat membrane caveolin-1 to caveolae and reduced transmigration. Here we report that cavin-1 expression reduces tumour size and metastasis of PC3 cells in vivo, using an orthotopic prostate cancer xenograft mouse model. To determine if cavin-1 acts by neutralizing oncogenic caveolin-1, we expressed cavin-1 in caveolin-1 negative LNCaP and 22Rv1 cells. While caveolin-1 over-expression increased anchorage-independent growth of LNCaP and 22Rv1 cells, cavin-1 over-expression had no effect. Furthermore, co-expression of cavin-1 in LNCaP+caveolin-1 cells reversed caveolin-1 effect. Taken together, these results suggest that caveolin-1 in prostate cancer is present outside of caveolae, and caveola formation by cavin-1 co-expression alters the oncogenic effect of non-caveolar caveolin-1 microdomains. Citation Format: Hyeongsun Moon, C Soon Lee, Sowmya Sharma, Kerry Inder, Debra Black, Kim-Anh Le Cao, Clay Winterford, Patrick Ling, the Australian Prostate Cancer BioResource, David Craik, Robert Parton, Pamela Russell, Michelle Hill. Cavin-1 alters oncogenic effects of caveolin-1 microdomains in prostate cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3040. doi:10.1158/1538-7445.AM2013-3040