Deficiency of the lysosomal protein CLN5 alters lysosomal function and movement

Batten disease is a devastating childhood rare neurodegenerative disease characterized by rapid deterioration of cognition and movement, leading to death within ten to thirty years of age. One of the thirteen Batten disease forms, CLN5 Batten disease, is caused by mutations in the CLN5 gene leading to motor deficits, mental deterioration, cognitive impairment, visual impairment, and epileptic seizures in children. A characteristic pathology in CLN5 Batten disease is the defects in lysosomes, leading to neuronal dysfunction. In this study, we aimed to investigate the lysosomal changes in CLN5-deficient human neurons. We used an induced pluripotent stem cell system, which generates pure human cortical-like glutamatergic neurons. Using CRISPRi, we inhibited the expression of CLN5 in human neurons. The CLN5-deficient human neurons showed neutralised lysosomal acidity and reduced lysosomal enzyme activity measured by microscopy and flow cytometry. Furthermore, the CLN5-deficient human neurons also showed impaired lysosomal movement, a phenotype that has never been reported in CLN5 Batten disease. Lysosomal trafficking is key to maintain local degradation of cellular wastes, especially in long neuronal projections and our results from the human neuronal model present a key finding to understand the underlying lysosomal pathology in neurodegenerative diseases

Aging germline stem cells in C. elegans

Failure to maintain stem cells with age is associated with conditions such as tissue degeneration and increased susceptibility to tissue damage. We use the C. elegans germline stem cell system as a model to study stem cell aging. This system combines a well-established model for aging with an accessible stem cell system, providing a unique opportunity to understand how aging influences stem cell dynamics. The germline stem/progenitor pool in in C. elegans becomes depleted over time. At the cellular level, aging influences both the size of the stem cell pool and the proliferation rate of stem cells. The flux of differentiated cells also affects how aging impacts the pool. This depletion is partially alleviated in mutants with reduced insulin/IGF-like signaling via inhibition of the transcription factor DAF-16/FOXO. In this role, DAF-16 does not act in the germ line, and its anatomical requirements are different from its previously described roles in larval germline proliferation, dauer control, and lifespan regulation. We found that DAF-16/FOXO is required in certain somatic cells in the proximal part of the reproductive system to regulate the stem cell pool. We also find that the degree to which various age-defying perturbations affect lifespan does not correlate with their effect on germline stem cell maintenance. We are investigating additional aspects of aging germline stem cells using this system.

Complementary In Vivo Approaches Reveal Quiescent Melanocytes in Anagen Outside the Hair Follicle Bulge

ABSTRACTMelanocyte stem cells (McSCs) are key components of the hair follicle (HF) stem cell system that are derived from neural crest during embryogenesis and are responsible for regeneration of differentiated melanocytes during successive HF cycles. Our previous research has shown presence of two subsets of phenotypically and functionally distinct McSCs exist in murine telogen HFs, CD34+ McSCs in the bulge/lower permanent portion (LPP) and CD34− McSCs in the secondary hair germ (SHG). Whether these subsets are maintained independently or exist in a developmental hierarchy is not yet known. Using Dct-H2BGFP mice, we analyzed the quiescent and proliferative properties of McSCs and melanocytes in anagen and telogen. We found unexpectedly that Kit+Nestin− quiescent melanocytes are maintained outside of the bulge/LPP region throughout anagen in addition to the Kit+Nestin+ quiescent melanocytes of the bulge/LPP. Both subpopulations express lower levels of melanocyte differentiation markers Mitf, Pax3, Dct, Tyrp1 and Tyr compared to differentiated melanocytes of the HF bulb/matrix. These results suggest that quiescent melanocytes localized in the outer root sheath, both in and below the bulge/LPP) retain the stem cell phenotype observed in quiescent McSCs during telogen. This finding has implications for maintenance of distinct subsets of McSCs throughout successive HF cycles.

Biology of the Caenorhabditis elegans Germline Stem Cell System

Stem cell systems regulate tissue development and maintenance. The germline stem cell system is essential for animal reproduction, controlling both the timing and number of progeny through its influence on gamete production. In this review, we first draw general comparisons to stem cell systems in other organisms, and then present our current understanding of the germline stem cell system in Caenorhabditis elegans. In contrast to stereotypic somatic development and cell number stasis of adult somatic cells in C. elegans, the germline stem cell system has a variable division pattern, and the system differs between larval development, early adult peak reproduction and age-related decline. We discuss the cell and developmental biology of the stem cell system and the Notch regulated genetic network that controls the key decision between the stem cell fate and meiotic development, as it occurs under optimal laboratory conditions in adult and larval stages. We then discuss alterations of the stem cell system in response to environmental perturbations and aging. A recurring distinction is between processes that control stem cell fate and those that control cell cycle regulation. C. elegans is a powerful model for understanding germline stem cells and stem cell biology.

WUSCHEL acts as an auxin response rheostat to maintain apical stem cells in Arabidopsis

To maintain the balance between long-term stem cell self-renewal and differentiation, dynamic signals need to be translated into spatially precise and temporally stable gene expression states. In the apical plant stem cell system, local accumulation of the small, highly mobile phytohormone auxin triggers differentiation while at the same time, pluripotent stem cells are maintained throughout the entire life-cycle. We find that stem cells are resistant to auxin mediated differentiation, but require low levels of signaling for their maintenance. We demonstrate that the WUSCHEL transcription factor confers this behavior by rheostatically controlling the auxin signaling and response pathway. Finally, we show that WUSCHEL acts via regulation of histone acetylation at target loci, including those with functions in the auxin pathway. Our results reveal an important mechanism that allows cells to differentially translate a potent and highly dynamic developmental signal into stable cell behavior with high spatial precision and temporal robustness.

A novel self-organizing embryonic stem cell system reveals signaling logic underlying the patterning of human ectoderm

SummaryDuring development, the ectoderm is patterned by a combination of BMP and WNT signaling. Research in model organisms has provided substantial insight, however, there are currently no systems to study this patterning in humans. Further, the complexity of neural plate border specification has made it difficult to transition from discovering the genes involved to deeper mechanistic understanding. Here, we develop an in vitro model of human ectodermal patterning, in which hESCs self-organize to form robust and quantitatively reproducible patterns corresponding to the dorsal-ventral axis of the embryo. Using this platform, we show that the duration of endogenous WNT signaling is a crucial control parameter, and that cells sense relative levels of BMP and WNT signaling in making fate decisions. These insights allowed us to develop an improved protocol for placodal differentiation. Thus, our platform is a powerful tool for studying human ectoderm patterning and for improving directed differentiation protocols.