Swathi Yadlapalli

Chromosome-specific nonrandom sister chromatid segregation during stem-cell division

Adult stem cells undergo asymmetric cell division to self-renew and give rise to differentiated cells that comprise mature tissue. Sister chromatids may be distinguished and segregated nonrandomly in asymmetrically dividing stem cells, although the underlying mechanism and the purpose it may serve remain elusive. Here we develop the CO-FISH (chromosome orientation fluorescence in situ hybridization) technique with single-chromosome resolution and show that sister chromatids of X and Y chromosomes, but not autosomes, are segregated nonrandomly during asymmetric divisions of Drosophila male germline stem cells. This provides the first direct evidence, to our knowledge, that two sister chromatids containing identical genetic information can be distinguished and segregated nonrandomly during asymmetric stem-cell divisions. We further show that the centrosome, SUN–KASH nuclear envelope proteins and Dnmt2 (also known as Mt2) are required for nonrandom sister chromatid segregation. Our data indicate that the information on X and Y chromosomes that enables nonrandom segregation is primed during gametogenesis in the parents. Moreover, we show that sister chromatid segregation is randomized in germline stem cell overproliferation and dedifferentiated germline stem cells. We propose that nonrandom sister chromatid segregation may serve to transmit distinct information carried on two sister chromatids to the daughters of asymmetrically dividing stem cells.

Drosophila male germline stem cells do not asymmetrically segregate chromosome strands

Adult stem cells continuously supply differentiated cells throughout the life of organisms. This increases the risk of replicative senescence or neoplastic transformation due to mutations that accumulate over many rounds of DNA replication. The immortal strand hypothesis proposes that stem cells reduce the accumulation of replication-induced mutations by retaining the older template DNA strands. Other models have also been proposed in which stem cells asymmetrically segregate chromosome strands for other reasons, such as retention of epigenetic memories. Recently, the idea has emerged that the mother centrosome, which is stereotypically retained within some asymmetrically dividing stem cells, might be utilized as a means of asymmetrically segregating chromosome strands. We have tested this hypothesis in germline stem cells (GSCs) from Drosophila melanogaster testis, which undergo asymmetric divisions marked by the asymmetric segregation of centrosomes and the acquisition of distinct daughter cell fates (stem cell self-renewal versus differentiation). Using 5-bromo-2-deoxyuridine labeling combined with direct visualization of GSC-gonialblast (differentiating daughter) pairs, we directly scored the outcome of chromosome strand segregation. Our data show that, in male GSCs in the Drosophila testis, chromosome strands are not asymmetrically segregated, despite asymmetrically segregating centrosomes. Our data demonstrate that asymmetric centrosome segregation in stem cells does not necessarily lead to asymmetric chromosome strand segregation.

DNA asymmetry in stem cells – immortal or mortal?

Summary The immortal strand hypothesis proposes that stem cells retain a template copy of genomic DNA (i.e. an ‘immortal strand’) to avoid replication-induced mutations. An alternative hypothesis suggests that certain cells segregate sister chromatids non-randomly to transmit distinct epigenetic information. However, this area of research has been highly controversial, with conflicting data even from the same cell types. Moreover, historically, the same term of ‘non-random sister chromatid segregation’ or ‘biased sister chromatid segregation’ has been used to indicate distinct biological processes, generating a confusion in the biological significance and potential mechanism of each phenomenon. Here, we discuss the models of non-random sister chromatid segregation, and we explore the strengths and limitations of the various techniques and experimental model systems used to study this question. We also describe our recent study on Drosophila male germline stem cells, where sister chromatids of X and Y chromosomes are segregated non-randomly during cell division. We aim to integrate the existing evidence to speculate on the underlying mechanisms and biological relevance of this long-standing observation on non-random sister chromatid segregation.

Circadian clock neurons constantly monitor environmental temperature to set sleep timing

Circadian clocks coordinate behaviour, physiology and metabolism with Earth’s diurnal cycle. These clocks entrain to both light and temperature cycles, and daily environmental temperature oscillations probably contribute to human sleep patterns. However, the neural mechanisms through which circadian clocks monitor environmental temperature and modulate behaviour remain poorly understood. Here we elucidate how the circadian clock neuron network of Drosophila melanogaster processes changes in environmental temperature. In vivo calcium-imaging techniques demonstrate that the posterior dorsal neurons 1 (DN1ps), which are a discrete subset of sleep-promoting clock neurons, constantly monitor modest changes in environmental temperature. We find that these neurons are acutely inhibited by heating and excited by cooling; this is an unexpected result when considering the strong correlation between temperature and light, and the fact that light excites clock neurons. We demonstrate that the DN1ps rely on peripheral thermoreceptors located in the chordotonal organs and the aristae. We also show that the DN1ps and their thermosensory inputs are required for the normal timing of sleep in the presence of naturalistic temperature cycles. These results identify the DN1ps as a major gateway for temperature sensation into the circadian neural network, which continuously integrates temperature changes to coordinate the timing of sleep and activity.

Spindle positioning in the stem cell niche

Stem cells are the source of differentiated cells that constitute tissues and organs. Two fundamental characteristics of stem cells are their abilities to self‐renew stem cell identity and to produce differentiated cells, the balance of which can be achieved by asymmetric stem cell division. Many stem cells have been shown to reside in a stem cell niche, the home of stem cells that regulates the stem cell behavior. Recent studies have revealed the critical contribution of cytoskeletons in achieving asymmetric stem cell division: mitotic spindles in dividing stem cells are often oriented with respect to the stem cell niche, which is supported by concerted actions of microtubule networks and components at the cell membrane such as adherens junctions, the actin cytoskeleton, and the extracellular matrix. In this article, we review the mechanism of stem cell spindle orientation, with emphasis on its relationship with the stem cell niche, and discuss how it contributes to tissue development and homeostasis. WIREs Dev Biol 2012, 1:215–230. doi: 10.1002/wdev.16

Parallelized, real-time, metabolic-rate measurements from individual Drosophila

Significant recent evidence suggests that metabolism is intricately linked to the regulation and dysfunction of complex cellular and physiological responses ranging from altered metabolic programs in cancers and aging to circadian rhythms and molecular clocks. While the metabolic pathways and their fundamental control mechanisms are well established, the precise cellular mechanisms underpinning, for example, enzymatic pathway control, substrate preferences or metabolic rates, remain far less certain. Comprehensive, continuous metabolic studies on model organisms, such as the fruit fly Drosophila melanogaster, may provide a critical tool for deciphering these complex physiological responses. Here, we describe the development of a high-resolution calorimeter, which combines sensitive thermometry with optical imaging to concurrently perform measurements of the metabolic rate of ten individual flies, in real-time, with ~100 nW resolution. Using this calorimeter we have measured the mass-specific metabolic rates of flies of different genotypes, ages, and flies fed with different diets. This powerful new approach enables systematic studies of the metabolic regulation related to cellular and physiological function and disease mechanisms.

How a brain keeps its cool

Temperature-sensing neurons in the Drosophila brain cooperate with the central circadian clock to help regulate body temperature.

Temperature Sensation and Integration in the Drosophila Circadian Clock

Circadian clocks are entrained by zeitgebers, environmental cues such as light and temperature that adapt living organisms to the physical rhythms of the earth. Although temperature has been shown to be a major zeitgeber and can entrain the circadian clock of Drosophila, the neural and molecular mechanisms by which circadian clocks respond to temperature remain poorly understood. In our work, we use in vivo calcium imaging to characterize the temperature response of clock neurons in Drosophila to temperature modulation. We show that a selective group of clock neurons responds to temperature changes and that dorsal neurons (DNs) are excited by cooling and inhibited by warming. We further investigated the physiological input pathway of temperature sensing into the circadian clock. We find that arista and chordotonal organs are both critical factors that contribute to the response of circadian neurons to temperature modulation. Our work reveals that clock neurons respond to temperature changes through multiple temperature input pathways, suggesting a complex network similar to the entrainment of circadian clocks by light input.

Past experience resets behavior: CaMK takes the heat.

How past experiences reshape behavior is not well understood. In this issue, two studies (Schild et al., 2014; Yu et al., 2014) dissected the molecular mechanisms underlying experience-dependent plasticity in thermosensory behavior. They show that Ca(2+)/calmodulin-dependent kinase I (CaMKI) regulates thermal preferences according to past experience.