| dc.description.abstract | Ever since Sydney Brenner introduced Caenorhabditis elegans in the early 60s as a
model system, this worm has contributed to many ground-breaking discoveries in
science. It has served as an excellent model to study animal development, metabolism,
longevity, neurobiology, and so on. Here, we have used C. elegans to understand the
genetic regulation of skin barrier function and longevity.
C. elegans dwells in an organically rich environment in the soil where it is exposed to
various bacterial toxins and toxic chemicals. Under such conditions, it becomes very
crucial for the worms to maintain the skin barrier integrity to prevent the diffusion of toxic
molecules into the body. However, the components of skin that provide a barrier against
diffusion of exogenous chemicals remain unknown. To identify genetic regulator(s) of
the skin barrier function, we screened 93 collagens of C. elegans for their requirement
in permeability barrier maintenance. We show that four collagens- DPY-7, DPY-8, DPY-9,
and DPY-10 are essential for maintaining the skin barrier function of C. elegans skin. Loss of
any of the 4 permeability-determining (PD) collagens leads to enhanced susceptibility of the
nematode to paraquat (PQ) and antihelminthic drugs, levamisole, and ivermectin. Upon
exposure to paraquat, PD collagen mutants accumulate more PQ, incur more damage, and
death despite the robust activation of antioxidant machinery. We show that the permeability
barrier maintained by PD collagens acts in parallel to FOXO transcription factor DAF-16 to
enhance survival of insulin-like receptor mutant, daf-2. In all, this study shows that PD
collagens regulate cuticle permeability by maintaining the structure of C. elegans skin and
thus provide protection against exogenous toxic molecules.
As PD collagens are essential for maintaining C. elegans cuticle structure, barrier function,
and survival against exogenous toxic molecules, they must be under tight transcriptional
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regulation. We screened 286 transcription factors expressed in hypodermis for their role in
permeability barrier maintenance. We show that BLMP-1, T26A8.4, and LIR-1, zinc finger
transcription factors, are important for maintaining the barrier function of the cuticle. blmp-1,
T26A8.4, and lir-1 RNAi animals phenocopy PD collagens for survival on PQ, ivermectin, and
levamisole. We further show that BLMP-1 and LIR-1 control cuticle permeability by regulating
the expression of PD collagens and collagen processing enzyme protein disulfide isomerase
pdi-2 in wild type animals. T26A8.4 RNAi altered expression of pdi-2 only. From this, we
conclude that BLMP-1 and LIR-1 are the major regulators of the permeability determining
collagens in C. elegans.
Cuticle barrier function study shows that maintenance of cuticle ultrastructure and function is
crucial for C. elegans survival. Recent studies show that C. elegans activates repair pathways
and immune response upon wounding suggesting C. elegans can sense damage to the skin.
We found that knockdown of PD collagens improved survival of C. elegans against osmotic
stress, thermal stress, and P. aeruginosa infection while causing susceptibility to PQ. PD
collagen animals also have a higher basal level of antioxidants, glyceraldehyde-5-phosphate
dehydrogenase, and antimicrobial peptides. This indicated that there must be a cuticle
structural and functional integrity surveillance system in the hypodermis. In microarray by
Rohlfing et al., 2010, Patch-like Receptor (PTR) family expression was altered in dpy-9 and
dpy-10 animals. By systematic analysis of 22 PTRs in dpy-9 animals for osmotic and hydrogen
peroxide, we found that enhanced resistance of PD collagen animals is dependent on a
specific receptor PTR-23. We found that PTR-23 works along with permeability determining
factor LIR-1 to regulate thermal resistance in dpy-10 animals. In all, this study shows that C.
elegans can perceive cuticle defects and trigger a stress response in a PTR-23 dependent
manner.
In nature, the only constant is change (Heraclitus, 500 BCE). To maintain an optimal life
span, an organism needs to sense the ever-changing environment and adapt
behaviorally and physiologically. This adaptation is coordinated by the nervous system
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of multicellular organisms. G-protein coupled receptors (GPCRs) are reported to sense
environmental cues and regulate behavior and physiology. A recent life span study in
Drosophila suggests GPCRs might regulate life span as well. In C. elegans, ablation of
olfactory neurons-AWA, AWB, and AWC influence longevity suggesting the importance
of olfaction and food sensing in the regulation of life span. We asked if specific olfactory
GPCRs regulate C. elegans life span? We show that C. elegans longevity is regulated by a
chemosensory GPCR STR-2, expressed in AWC and ASI amphid sensory neurons. This
neuronal receptor controls lipid droplet homeostasis in the intestine. We show that STR-2
specifically regulates the expression of delta-9 desaturases, fat-5, fat-6, and fat-7 responsible
for the production of monosaturated fatty acids (MUFA), an indicator of health and longevity,
and diacylglycerol acyltransferase dgat-2. DGAT-2 catalyzes the final step of triglyceride
synthesis i.e., transfer of a long-chain fatty acyl-CoA to diacylglycerol and therefore required
for lipid droplet synthesis. Rescue of stored fat levels of GPCR mutant animals to wild type
levels, with supplementation of the diet with a low concentration of glucose, rescues its life
span phenotype. In all, we show that neuronal STR-2 GPCR facilitates metabolic adaptation
to maintain the optimal life span at higher temperatures in C. elegans.
In summary, we show that the optimal survival of C. elegans is dependent on intact cuticle
barrier function and their ability to adapt to the constantly changing environment. The study of
C. elegans barrier function highlights the importance of maintaining skin barricade by specific
collagens. In mammalian skin, the stratum corneum of the skin act as an impermeable barrier
against exogenous toxic molecules. Skin barrier defect has been associated with several
diseases in humans such as Gaucher disease, atopic dermatitis, psoriasis, etc, and often
associated with inflammation. C. elegans PD collagen mutants can serve as a tool to study
toxicity or infiltration of high molecular weight toxic molecules such as commonly used
herbicides and pesticides. C. elegans collagen skin could also be used to address permeability
barrier function loss due to injury and, its impact on inflammation and wound healing
responses. The study of life span regulation by a neuronal GPCR STR-2 suggests that GPCRs
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in the nervous system and other tissues could play an important role in determining the life
span of an organism by regulating metabolic adaptation to environmental stimuli. Identifying
the environmental cues, neural circuits, and genetic regulators could help us understand the
mechanisms of aging better. This can help us design proper prognosis and treatment for age-associated diseases | en_US |
