Autophagy regulator BECN1 suppresses mammary tumorigenesis driven by WNT1 activation and following parity

Earlier studies reported allelic deletion of the essential autophagy regulator BECN1 in breast cancers implicating BECN1 loss, and likely defective autophagy, in tumorigenesis. Recent studies have questioned the tumor suppressive role of autophagy, as autophagy-related gene (Atg) defects generally suppress tumorigenesis in well-characterized mouse tumor models. We now report that, while it delays or does not alter mammary tumorigenesis driven by Palb2 loss or ERBB2 and PyMT overexpression, monoallelic Becn1 loss promotes mammary tumor development in 2 specific contexts, namely following parity and in association with wingless-type MMTV integration site family, member 1 (WNT1) activation. Our studies demonstrate that Becn1 heterozygosity, which results in immature mammary epithelial cell expansion and aberrant TNFRSF11A/TNR11/RANK (tumor necrosis factor receptor superfamily, member 11a, NFKB activator) signaling, promotes mammary tumorigenesis in multiparous FVB/N mice and in cooperation with the progenitor cell-transforming WNT1 oncogene. Similar to our Becn1^+/?;MMTV-Wnt1 mouse model, low BECN1 expression and an activated WNT pathway gene signature correlate with the triple-negative subtype, TNFRSF11A axis activation and poor prognosis in human breast cancers. Our results suggest that BECN1 may have nonautophagy-related roles in mammary development, provide insight in the seemingly paradoxical roles of BECN1 in tumorigenesis, and constitute the basis for further studies on the pathophysiology and treatment of clinically aggressive triple negative breast cancers (TNBCs).     

APE1 Incision Activity at Abasic Sites in Tandem Repeat Sequences

Repetitive DNA sequences, such as those present in microsatellites and minisatellites, telomeres, and trinucleotide repeats (linked to fragile X syndrome, Huntington disease, etc.), account for nearly 30% of the human genome. These domains exhibit enhanced susceptibility to oxidative attack to yield base modifications, strand breaks, and abasic sites; have a propensity to adopt non-canonical DNA forms modulated by the positions of the lesions; and, when not properly processed, can contribute to genome instability that underlies aging and disease development. Knowledge on the repair efficiencies of DNA damage within such repetitive sequences is therefore crucial for understanding the impact of such domains on genomic integrity. In the present study, using strategically designed oligonucleotide substrates, we determined the ability of human apurinic/apyrimidinic endonuclease 1 (APE1) to cleave at apurinic/apyrimidinic (AP) sites in a collection of tandem DNA repeat landscapes involving telomeric and CAG/CTG repeat sequences. Our studies reveal the differential influence of domain sequence, conformation, and AP site location/relative positioning on the efficiency of APE1 binding and strand incision. Intriguingly, our data demonstrate that APE1 endonuclease efficiency correlates with the thermodynamic stability of the DNA substrate. We discuss how these results have both predictive and mechanistic consequences for understanding the success and failure of repair protein activity associated with such oxidatively sensitive, conformationally plastic/dynamic repetitive DNA domains.     

Autophagy regulator BECN1 suppresses mammary tumorigenesis driven by WNT1 activation and following parity

Earlier studies reported allelic deletion of the essential autophagy regulator BECN1 in breast cancers implicating BECN1 loss, and likely defective autophagy, in tumorigenesis. Recent studies have questioned the tumor suppressive role of autophagy, as autophagy-related gene (Atg) defects generally suppress tumorigenesis in well-characterized mouse tumor models. We now report that, while it delays or does not alter mammary tumorigenesis driven by Palb2 loss or ERBB2 and PyMT overexpression, monoallelic Becn1 loss promotes mammary tumor development in 2 specific contexts, namely following parity and in association with wingless-type MMTV integration site family, member 1 (WNT1) activation. Our studies demonstrate that Becn1 heterozygosity, which results in immature mammary epithelial cell expansion and aberrant TNFRSF11A/TNR11/RANK (tumor necrosis factor receptor superfamily, member 11a, NFKB activator) signaling, promotes mammary tumorigenesis in multiparous FVB/N mice and in cooperation with the progenitor cell-transforming WNT1 oncogene. Similar to our Becn1^+/−;MMTV-Wnt1 mouse model, low BECN1 expression and an activated WNT pathway gene signature correlate with the triple-negative subtype, TNFRSF11A axis activation and poor prognosis in human breast cancers. Our results suggest that BECN1 may have nonautophagy-related roles in mammary development, provide insight in the seemingly paradoxical roles of BECN1 in tumorigenesis, and constitute the basis for further studies on the pathophysiology and treatment of clinically aggressive triple negative breast cancers (TNBCs).     

Development,cross-species/genera transferability of novel EST-SSR markers and their utility in revealing population structure and genetic diversity in sugarcane

Sugarcane (Saccharum spp. hybrid) with complex polyploid genome requires a large number of informative DNA markers for various applications in genetics and breeding. Despite the great advances in genomic technology, it is observed in several crop species, especially in sugarcane, the availability of molecular tools such as microsatellite markers are limited. Now-a-days EST-SSR markers are preferred to genomic SSR (gSSR) as they represent only the functional part of the genome, which can be easily associated with desired trait. The present study was taken up with a new set of 351 EST-SSRs developed from the 4085 non redundant EST sequences of two Indian sugarcane cultivars. Among these EST-SSRs, TNR containing motifs were predominant with a frequency of 51.6%. Thirty percent EST-SSRs showed homology with annotated protein. A high frequency of SSRs was found in the 5′UTR and in the ORF (about 27%) and a low frequency was observed in the 3′UTR (about 8%). Two hundred twenty-seven EST-SSRs were evaluated, in sugarcane, allied genera of sugarcane and cereals, and 134 of these have revealed polymorphism with a range of PIC value 0.12 to 0.99. The cross transferability rate ranged from 87.0% to 93.4% in Saccharum complex, 80.0% to 87.0% in allied genera, and 76.0% to 80.0% in cereals. Cloning and sequencing of EST-SSR size variant amplicons revealed that the variation in the number of repeat-units was the main source of EST-SSR fragment polymorphism. When 124 sugarcane accessions were analyzed for population structure using model-based approach, seven genetically distinct groups or admixtures thereof were observed in sugarcane. Results of principal coordinate analysis or UPGMA to evaluate genetic relationships delineated also the 124 accessions into seven groups. Thus, a high level of polymorphism adequate genetic diversity and population structure assayed with the EST-SSR markers not only suggested their utility in various applications in genetics and genomics in sugarcane but also enriched the microsatellite marker resources in sugarcane.     

Overexpression of the single-stranded DNA-binding protein (SSB) stabilises CAG•CTG triplet repeats in an orientation dependent manner

The stability and deletion-size-distribution profiles of leading strand (CAG)₇₅ and (CTG)₁₃₇ trinucleotide repeat arrays inserted in the Escherichia coli chromosome were investigated upon overexpression of the single-stranded DNA-binding protein (SSB) and in mutant strains deficient for the SbcCD (Rad51/Mre11) nuclease. SSB overexpression increases the stability of the (CAG)₇₅ repeat array and leads to a loss of the bias towards large deletions for the same array. Furthermore, the absence of SbcCD leads to a reduction in the number of large deletions in strains containing the (CTG)₁₃₇ repeat array.     

Fen1 does not control somatic hypermutability of the (CTG)(n)*(CAG)(n) repeat in a knock-in mouse model for DM1

The mechanism of trinucleotide repeat expansion, an important cause of neuromuscular and neurodegenerative diseases, is poorly understood. We report here on the study of the role of flap endonuclease 1 (Fen1), a structure-specific nuclease with both 5' flap endonuclease and 5'-3' exonuclease activity, in the somatic hypermutability of the (CTG)(n)*(CAG)(n) repeat of the DMPK gene in a mouse model for myotonic dystrophy type 1 (DM1). By intercrossing mice with Fen1 deficiency with transgenics with a DM1 (CTG)(n)*(CAG)(n) repeat (where 104n110), we demonstrate that Fen1 is not essential for faithful maintenance of this repeat in early embryonic cleavage divisions until the blastocyst stage. Additionally, we found that the frequency of somatic DM1 (CTG)(n)*(CAG)(n) repeat instability was essentially unaltered in mice with Fen1 haploinsufficiency up to 1.5 years of age. Based on these findings, we propose that Fen1, despite its role in DNA repair and replication, is not primarily involved in maintaining stability at the DM1 locus.     

A Survey of Attitudes of Local Citizens of a Residential Area Toward Urban Stray Cats in Japan

This study surveyed the attitudes of local residents living in an urban area in Japan toward stray cats. An anonymous questionnaire asked local residents (359 houses) about their attitudes toward stray cats. Responses were received from126 houses (35%). Answers about nuisance, respondents' actions, and actions to be taken with regard to stray cats did not differ by place or type of residence of respondents. More than one third (36.7 ± 16.6%) of the respondents answered that the “bad smell of the feces and urine” was a nuisance. Respondents who lived in detached houses tended to like cats compared with those who lived in condominiums. Respondents who liked cats took care of cats more frequently, whereas those who disliked cats chased cats away and prevented their intrusion into their houses and land. However, it is noteworthy that one third or more (minimum value: 37.8%) of respondents of all kinds answered that neutering is one effective way to suppress the population of stray cats.