high-throughput-screening-microbial-isolates-with-impact-on

High-Throughput Screening of Microbial Isolates with Impact on <em>Caenorhabditis elegans</em> Health

With its small size, short lifespan, and easy genetics, Caenorhabditis elegans offers a convenient platform to study the impact of microbial isolates on ...

Lancaster University

All living cells display a rapid molecular response to adverse environmental conditions, and the heat shock protein family reflects one such example. Hence, failing to activate heat shock proteins can impair the cellular response. In the present study, we evaluated whether the loss of different isoforms of heat shock protein (hsp) genes in Caenorhabditis elegans would affect their vulnerability to Manganese (Mn) toxicity.

Involvement of heat shock proteins on Mn-induced toxicity in Caenorhabditis elegans.

In C. elegans, the skn-1 gene encodes a transcription factor that resembles mammalian Nrf2 and activates a detoxification response. skn-1 promotes resistance to oxidative stress (Oxr) and also increases lifespan, and it has been suggested that the former causes the latter, consistent with the theory that oxidative damage causes aging. Here, we report that effects of SKN-1 on Oxr and longevity can be dissociated. We also establish that skn-1 expression can be activated by the DAF-16/FoxO transcription factor, another central regulator of growth, metabolism, and aging. Notably, skn-1 is required for Oxr but not increased lifespan resulting from over-expression of DAF-16; concomitantly, DAF-16 over-expression rescues the short lifespan of skn-1 mutants but not their hypersensitivity to oxidative stress. These results suggest that SKN-1 promotes longevity by a mechanism other than protection against oxidative damage.

The SKN-1/Nrf2 transcription factor can protect against oxidative stress and increase lifespan in C. elegans by distinct mechanisms.

Organismal death is a process of systemic collapse whose mechanisms are less well understood than those of cell death. We previously reported that death in C. elegans is accompanied by a calcium-propagated wave of intestinal necrosis, marked by a wave of blue autofluorescence (death fluorescence). Here, we describe another feature of organismal death, a wave of body wall muscle contraction, or death contraction (DC). This phenomenon is accompanied by a wave of intramuscular Ca release and, subsequently, of intestinal necrosis. Correlation of directions of the DC and intestinal necrosis waves implies coupling of these death processes. Long-lived insulin/IGF-1-signaling mutants show reduced DC and delayed intestinal necrosis, suggesting possible resistance to organismal death. DC resembles mammalian rigor mortis, a postmortem necrosis-related process in which Ca influx promotes muscle hyper-contraction. In contrast to mammals, DC is an early rather than a late event in C. elegans organismal death. VIDEO ABSTRACT.

Coupling of Rigor Mortis and Intestinal Necrosis during C. elegans Organismal Death.

A long-standing belief is that aging (senescence) is the result of stochastic damage accumulation. Alternatively, senescent pathology may also result from late-life, wild-type gene action (i.e., antagonistic pleiotropy, as argued by Williams) leading to non-adaptive run-on of developmental programs (or ) (as suggested more recently by Blagosklonny). In this study, we use existing and new data to show how uterine tumors, a prominent form of senescent pathology in the nematode , likely result from quasi-programs. Such tumors develop from unfertilized oocytes which enter the uterus and become hypertrophic and replete with endoreduplicated chromatin masses. Tumor formation begins with ovulation of unfertilized oocytes immediately after exhaustion of sperm stocks. We show that the timing of this transition between program and quasi-program (i.e., the onset of senescence), and the onset of tumor formation, depends upon the timing of sperm depletion. We identify homology between uterine tumors and mammalian ovarian teratomas, which both develop from oocytes that fail to mature after meiosis I. In teratomas, futile activation of developmental programs leads to the formation of differentiated structures within the tumor. We report that older uterine tumors express markers of later embryogenesis, consistent with teratoma-like activation of developmental programs. We also present evidence of coupling of distal gonad atrophy to oocyte hypertrophy. This study shows how the Williams Blagosklonny model can provide a mechanistic explanation of this component of  aging. It also suggests etiological similarity between teratoma and some forms of senescent pathology, insofar as both are caused by quasi-programs.

A parthenogenetic quasi-program causes teratoma-like tumors during aging in wild-type .

Aging (senescence) is characterized by the development of numerous pathologies, some of which limit lifespan. Key to understanding aging is discovery of the mechanisms (etiologies) that cause senescent pathology. In C. elegans, a major senescent pathology of unknown etiology is atrophy of its principal metabolic organ, the intestine. Here we identify a cause of not only this pathology but also of yolky lipid accumulation and redistribution (a form of senescent obesity): autophagy-mediated conversion of intestinal biomass into yolk. Inhibiting intestinal autophagy or vitellogenesis rescues both visceral pathologies and can also extend lifespan. This defines a disease syndrome leading to multimorbidity and contributing to late-life mortality. Activation of gut-to-yolk biomass conversion by insulin/IGF-1 signaling (IIS) promotes reproduction and senescence. This illustrates how major, IIS-promoted senescent pathologies in C. elegans can originate not from damage accumulation but from direct effects of futile, continued action of a wild-type biological program (vitellogenesis).

C. elegans Eats Its Own Intestine to Make Yolk Leading to Multiple Senescent Pathologies.

A plethora of studies over several decades has demonstrated the importance of autophagy in aging and age-related neurodegenerative disease. The role of autophagy in damage clearance and cell survival is well established, and supports a prevailing view that increasing autophagic activity can be broadly beneficial, and could form the basis of anti-aging interventions. However, macroautophagy/autophagy also promotes some elements of senescence. For example, in C. elegans hermaphrodites it facilitates conversion of intestinal biomass into yolk, leading to sex-specific gut atrophy and senescent steatosis.

Autophagy promotes visceral aging in wild-type C. elegans.

Caenorhabditis elegans is an excellent model for high-throughput experimental approaches but lacks an automated means to pinpoint time of death during survival assays over a short time frame, that is, easy to implement, highly scalable, robust, and versatile. Here, we describe an automated, label-free, high-throughput method using death-associated fluorescence to monitor nematode population survival (dubbed LFASS for label-free automated survival scoring), which we apply to severe stress and infection resistance assays. We demonstrate its use to define correlations between age, longevity, and severe stress resistance, and its applicability to parasitic nematodes. The use of LFASS to assess the effects of aging on susceptibility to severe stress revealed an unexpected increase in stress resistance with advancing age, which was largely autophagy-dependent. Correlation analysis further revealed that while severe thermal stress resistance positively correlates with lifespan, severe oxidative stress resistance does not. This supports the view that temperature-sensitive protein-handling processes more than redox homeostasis underpin aging in C. elegans. That the ages of peak resistance to infection, severe oxidative stress, heat shock, and milder stressors differ markedly suggests that stress resistance and health span do not show a simple correspondence in C. elegans.

New label-free automated survival assays reveal unexpected stress resistance patterns during C. elegans aging.

Alzheimer's disease (AD) is the leading form of dementia but lacks curative treatments. Current understanding of AD aetiology attributes the development of the disease to the misfolding of two proteins; amyloid-β (Aβ) and hyperphosphorylated tau, with their pathological accumulation leading to concomitant oxidative stress, neuroinflammation, and neuronal death. These processes are regulated at multiple levels to maintain homeostasis and avert disease. However, many of the relevant regulatory proteins appear to be downregulated in the AD-afflicted brain. Enhancement/restoration of these 'protective' proteins, therefore, represents an attractive therapeutic avenue. Gene therapy is a desirable means of achieving this because it is not associated with the side-effects linked to systemic protein administration, and sustained protein expression virtually eliminates compliance issues. The current article represents a focused and succinct review of the better established 'protective' protein targets for gene therapy enhancement/restoration rather than being designed as an exhaustive review incorporating less validated protein subjects. In addition, we will discuss how the risks associated with uncontrolled or irreversible gene expression might be mitigated through combining neuronal-specific promoters, inducible expression systems and localised injections. Whilst many of the gene therapy targets reviewed herein are yet to enter clinical trials, preclinical testing has thus far demonstrated encouraging potential for the gene therapy-based treatment of AD.

Gene therapy-mediated enhancement of protective protein expression for the treatment of Alzheimer's disease.

The Alzheimer's disease (AD)-associated amyloid-beta protein precursor (AβPP) can be cleaved by β-site AβPP cleaving enzyme 1 (BACE1) and the γ-secretase complex to yield neurotoxic amyloid-β (Aβ) peptides. However, AβPP can also be cleaved in a 'non-amyloidogenic' manner either by α-secretase to produce soluble AβPP alpha (sAβPPα) (a fragment with neuroprotective/neurogenic functions) or through alternative BACE1-mediated 'beta prime' activity yielding soluble AβPP beta prime (sAβPPβ').

BACE1 Overexpression Reduces SH-SY5Y Cell Viability Through a Mechanism Distinct from Amyloid-β Peptide Accumulation: Beta Prime-Mediated Competitive Depletion of sAβPPα.

Article	Year
Involvement of heat shock proteins on Mn-induced toxicity in Caenorhabditis elegans. BMC pharmacology & toxicology\| PubMed ID: 27802836	2016
The SKN-1/Nrf2 transcription factor can protect against oxidative stress and increase lifespan in C. elegans by distinct mechanisms. Aging cell\| PubMed ID: 28612944	2017
Coupling of Rigor Mortis and Intestinal Necrosis during C. elegans Organismal Death. Cell reports\| PubMed ID: 29514100	2018
A parthenogenetic quasi-program causes teratoma-like tumors during aging in wild-type . NPJ aging and mechanisms of disease\| PubMed ID: 29928508	2018
C. elegans Eats Its Own Intestine to Make Yolk Leading to Multiple Senescent Pathologies. Current biology : CB\| PubMed ID: 30100339	2018
Autophagy promotes visceral aging in wild-type C. elegans. Autophagy\| PubMed ID: 30700231	2019
New label-free automated survival assays reveal unexpected stress resistance patterns during C. elegans aging. Aging cell\| PubMed ID: 31309734	2019
Gene therapy-mediated enhancement of protective protein expression for the treatment of Alzheimer's disease. Brain research\| PubMed ID: 33422539	2021
BACE1 Overexpression Reduces SH-SY5Y Cell Viability Through a Mechanism Distinct from Amyloid-β Peptide Accumulation: Beta Prime-Mediated Competitive Depletion of sAβPPα. Journal of Alzheimer's disease : JAD\| PubMed ID: 35180123	2022

Alexandre Benedetto

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